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Molecular Biology
ofAssemblies
and Machines
Molecular Biology
ofAssemblies
and Machines

Alasdair C. Steven
Wolfgang Baumeister
Louise N. Johnson
Richard N. Perham
Garland Science
Vice President: Denise Schanck
Senior Editor: Summers Scholl
Editorial Assistants: Michael Roberts and William Sudry
Developmental Editor: Mary Purton
Production Editors and Layout: Georgina Lucas and
EJ Publishing Services
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Cover Design: Matthew McClements, Blink Studio, Ltd
Copyeditors: Bruce Goatly and Sally Livitt
Proofreader: Sally Huish
Indexer: Liza Furnival, Medical Indexing Limited
Permissions Coordinator: Sheri Gilbert
© 2016 by Garland Science, Taylor & Francis Group, LLC
This book contains information obtained from authentic
and highly regarded sources. Every effort has been made
to trace copyright holders and to obtain their permission
for the use of copyright material. Reprinted material is
quoted with permission, and sources are indicated. A
wide variety of references are listed. Reasonable efforts
have been made to publish reliable data and information,
but the author and the publisher cannot assume
responsibility for the validity of all materials or for the
consequences of their use.
All rights reserved. No part of this publication may be
reproduced, stored in a retrieval system or transmitted
in any form or by any means—graphic, electronic, or
mechanical, including photocopying, recording, taping,
or information storage and retrieval systems—without
permission of the copyright holder.
ISBN 978-0-8153-4166-6
About the Authors
Alasdair C. Steven trained as a theoretical physicist (PhD Cambridge), before
migrating into biology as a postdoctoral fellow at the Biozentrum, University of
Basel. As a principal investigator, his research is aimed at elucidating structure–
function–assembly–dynamics relationships of macromolecular complexes, such
as viruses, energy-dependent proteases, and amyloid filaments. He has developed
novel methods for image reconstruction and analysis of electron micrographs,
and ‘hybrid approaches’ in which cryo-electron microscopy is systematically
combined with other experimental methods.
Wolfgang Baumeister received his PhD in biophysics from the University of
Düsseldorf where he remained, apart from a year as Heisenberg Scholar at
the University of Cambridge, before moving to the Max-Planck-Institute of
Biochemistry in Martinsried. There he is head of the Department of Structural
Molecular Biology and an Honorary Professor of Physics at the Technical
University of Munich. Baumeister’s laboratory pioneered the development of
cryo-electron tomography enabling structural biology studies in situ. His work
has shaped the understanding of the structure and function of the cellular
machinery of protein degradation, most notably the proteasome.
Dame Louise N. Johnson was a leader in X-ray crystallography who contributed
to solving the first (lysozyme) and second (RNase) enzymes to be described by
this technique. She received her PhD from University College London, spent
a postdoctoral year at Yale, and made her career at the University of Oxford,
where she was Professor of Structural Biology from 1990–2007. She also served
as Director of Life Sciences at Diamond Light Source. Johnson’s laboratory
determined the structures of protein kinases, regulatory proteins of the cell cycle,
and glycogen metabolism—yielding insights as to the origin of diseases and drug
design.
Richard N. Perham was a protein biochemist with broad-ranging interests.
After national service in the Royal Navy he embarked on an academic career
(PhD from Cambridge; postdoctoral fellowship at Yale). He then returned to
Cambridge eventually becoming Chair of the Department of Biochemistry and
Master of St John’s College. Among the systems he studied were tobacco mosaic
virus assembly, bacteriorhodopsin, and multienzyme complexes including
pyruvate dehydrogenase. Perham also made influential contributions to synthetic
biology and commercial applications of phage display.

Library of Congress Cataloging-in-Publication Data

Names: Steven, Alasdair C., author. | Baumeister, W. (Wolfgang), 1946- ,
author. | Johnson, L. N., author. | Perham, R. N., author.
Title: Molecular biology of assemblies and machines / Alasdair C. Steven,
Wolfgang Baumeister, Louise N. Johnson, Richard N. Perham.
Description: New York, NY : Garland Science, [2016] | Includes
bibliographical references.
Identifiers: LCCN 2015042845 | ISBN 9780815341666 (alk. paper)
Subjects: | MESH: Macromolecular Substances--metabolism. | Biochemical
Processes. | Biophysical Processes. | Cell Physiological Processes. |
Macromolecular Substances--chemistry.
Classification: LCC QP171 | NLM QU 45 | DDC 572/.4--dc23
LC record available at http://lccn.loc.gov/2015042845

Published by Garland Science, Taylor & Francis Group, LLC, an informa business,
711 Third Avenue, New York, NY 10017, USA, and 3 Park Square, Milton Park, Abingdon, OX14 4RN, UK.

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FOREWORD

Structural biology has become a popular subject. Scientific journals use beautiful pictures
of protein folds to decorate their covers. Even advertisers in the popular press resort to
pictures of DNA or alpha helices to demonstrate the good scientific pedigree of their
products. In Molecular Biology of Assemblies and Machines, biology is explained in terms
of these well-known structural motifs. The book covers almost every basic biological topic
at a level that will allow the inquisitive reader to quickly absorb the fundamentals in terms
of the currently available structural information. The four authors bring together extensive
knowledge of biology, chemistry, biochemistry, physics, and mathematics from their diverse
backgrounds. Together they emphasize the mechanistic basics of biology and provide
insights that are often missing in books with more restricted interests. These will stimulate
the reader to look further for greater detail elsewhere.
This book is by no means the first one that views biology from a structural point of view.
One of the earliest books of this genre was written by Dickerson and Geis in 1969. Others
followed, including those by Schulz and Schirmer (1980), Brändén and Tooze (1991),
and Perutz (1992). But these books tend to concentrate on specific biological topics that
have seen significant structural achievements. The present book reverses this approach by
surveying the complete biological landscape in terms of the functions required by a living
cell. Then, currently available structural information is used to illuminate the mechanisms
of the biological machines that are required to maintain life.
The first protein structure was determined in the 1950s for sperm whale myoglobin, which
has a molecular mass of 17 kDa. As technology advanced, it became possible to solve much
larger complexes. For instance, some 20 years later, viruses with icosahedral symmetry
built of 180 identical subunits, each as big or bigger than myoglobin, were first described
and more recently, much larger viruses have been described in comparable detail. Many
of the molecular machines covered in this book have multiple copies of many different
subunits—as for instance the well-known and much studied bacteriophage T4. This is to be
expected as many biological functions are complex and require the coordinated efforts of
different specialized subunits. The larger the complex (for instance, a ribosome), the more
multifaceted will be the functions required of it; also, the less likely it is that every assembled
unit will have the exact same structure. Variants can arise from functionally relevant
conformational changes, but also from mistakes occurring during assembly, much as they do
in the assembling of houses from bricks. It is often difficult to exactly repeat an experiment
performed with biological macromolecules; indeed that is why the science of biology is
often considered to be an inexact science. Thus the wonderful structural tools developed
for relatively small, homogeneous molecules like myoglobin fade into obsolescence as
investigations are extended to larger assemblies that individually have small differences but
yet function similarly. Eventually structural investigations at near-atomic resolution must
address whole cells. In this case the assumption of structural uniformity that has brought us
to the present state of knowledge will need to be completely discarded. This book is being
published at a time when major changes are happening in the technology and direction of
structural biology, whereby objects of interest each have their own individual structure and
yet are functionally similar.

Michael G. Rossmann
PREFACE

We were motivated to undertake this book project based on the dawning realization—already
evident more than 10 years ago—that virtually all major biological activities are carried out not
by individual macromolecules, but by assemblies thereof, ranging from several to very many
components. These assemblies can be either transient or long-lived and they may integrate
multiple reactions or interconnect signaling pathways. This is the realm of mesobiology. Among
the assemblies, we distinguish an important subset with unmistakably machine-like properties,
while others can be viewed as ‘smart biomaterials.’ It is assemblies that generate energy; replicate
and repair DNA (the master-molecules of life); transcribe it into RNA; synthesize proteins and
assist in their folding; dispose of proteins that are defective or no longer needed; control the cell
cycle; communicate within and between cells; transport material between compartments; and
perform many other vital functions. Our goal, then, became to give a systematic account of their
structures and provide a mechanistic understanding in the light of the structures. It was clear at
the outset that this was a moving target: the extent to which they are understood varies widely
from system to system, and new assemblies are still being discovered on a regular basis. With
that caveat, we are as up-to-date as possible and our emphasis has been on structural principles
rather than an encyclopedic compilation.
This textbook is intended to be accessible to advanced undergraduates and graduate students
throughout the life sciences, and should also appeal to constituencies in physics, chemistry, and
engineering—especially in the machine-like aspects. Some familiarity with the basics of protein
structure and cell biology and in biochemistry and genetics will be helpful, but Chapter 1 of
this volume is intended to provide some background and set the stage. The subsequent chapters
should then enable the reader to move further into the original literature on given topics.
While there is some emphasis on the assemblies of eukaryotic cells, those of bacteria, archaea,
and even viruses, have not been neglected. In many cases, the assemblies of prokaryotes
afford valuable insights as simpler and experimentally tractable versions of their eukaryotic
counterparts.
We also mention, where possible, how many diseases have their roots in malfunctions of
assemblies or in mutations in their components. Aberrant assembly is the basis of sickle cell
disease; here a single mutation in hemoglobin causes filaments to assemble that distort the red
blood cells, impeding the circulation and thus eliciting the symptoms of this disease. More
recently, it has been recognized that aberrant assembly of another kind of protein filament—
amyloids, which are misfolded insoluble aggregates of some cellular proteins—is deeply
implicated in neurodegenerative diseases. One of these, Alzheimer’s, is emerging as the major
medical challenge of our time. Mechanistically in another direction, aberrant phosphorylation
by mutated kinases and their impact on signaling underlie many cancers.
A wealth of relevant information—graphic and otherwise—is already available on the Internet.
We see these resources as complementary to rather than as an alternative to what our textbook
has to offer. It will also be useful for the reader to have some familiarity with a user-friendly
molecular graphics program—for example, Pymol (www.pymol.org), Proteopedia (proteopedia.
org), or Chimera (http://www.cgl.ucsf.edu/)—to display, manipulate, and otherwise familiarize
themselves with the three-dimensional organization of assemblies of interest.
On didactics, we envisage that this textbook can provide a basis for many different college courses,
particularly those focused on the structures and functions of biological macromolecules. Possible
courses include but are not limited to: the biophysics or biochemistry of macromolecules;
macromolecular structure, function, and dynamics; macromolecular assemblies and/or
machines. Given the density of material covered, the text can also be tailored to suit concentrated
advanced courses. A one-semester course might typically cover Chapter 1 plus three or four
complementary and more focused chapters. For instance, a course on structural genetics might
include Chapters 2 (chromatin), 3 (DNA replication), and 4 (DNA repair); one addressing
biomolecular synthesis might be based on Chapters 5 (transcription), 6 (protein synthesis),
7 (protein degradation), and 8 (mechanisms of self-assembly); communications, motility
and transport might be covered with Chapters 10, 14, and 16; the enzymology and catalytic
viii Preface

properties of multiprotein complexes could be based on Chapters 9, 15, and 17; and the
assemblies that govern some cellular behaviors might go for Chapters 12, 13, and 17. Many
other combinations of chapters should also work well together.
As Lord Rutherford famously put it “All science is either physics or stamp-collecting.” In fact,
we may now detect a convergence between physics and structural biology. The synchrotron
beam lines and electron microscopes that made possible the structure determinations that
are defining mesobiology had their origins in physics. Indeed as this discipline moves closer
to maturity it may be seen as evolving toward a physics of complex systems. However,
science—like beauty—is in the eye of the beholder. The structures of macromolecular
assemblies such as the ribosome, the proteasome, and complex viruses stand out in
their visual beauty and high value—just like the ‘Penny Blacks’ and ‘Cape of Good Hope
triangulars’ in the eyes of philatelists.
Finally, we close by acknowledging the vital roles played by our coauthors, Louise Johnson
and Richard Perham, who have not lived to see completion of our project. It has been a
joy and a privilege to work with them. We owe much to Garland’s indefatigable Summers
Scholl, to Mary Purton for expert developmental editing, and to Nigel Orme for elegant
and insightful artwork. We thank Umar Salam for his support and help during the book’s
development. We affectionately thank Ray Steven, Gudrun Baumeister, and Nancy Lane
Perham for encouragement and support throughout this project.

RESOURCES FOR INSTRUCTORS AND STUDENTS

The teaching and learning resources for instructors and students are available online. The
instructor’s resources on the Garland Science website are password-protected and available
only to adopting instructors. The student resources on the Garland Science website are
available to everyone. We hope these resources will enhance student learning and make it
easier for instructors to prepare dynamic lectures and activities for the classroom.

Instructor Resources
Instructor Resources are available on the Garland Science Instructor Resource Center,
located at www.garlandscience.com/instructors. The website provides access not only to the
teaching resources for this book but also to all other Garland Science textbooks. Adopting
instructors can obtain access to the site from their sales representative or by emailing
science@garland.com.

Art of Molecular Biology of Assemblies and Machines

The images from the book are available in two convenient formats: PowerPoint® and JPEG.
They have been optimized for display on a computer. Figures are searchable by figure
number, by figure name, or by keywords used in the figure legend from the book.

Student Resources
The resources for students are available on the Molecular Biology of Assemblies and Machines
Student Website, located at www.garlandscience.com and selecting the ‘Student’ tab.

Flashcards
Each chapter contains flashcards, built into the student website, that allow students to
review key terms from the text.

Glossary
The comprehensive glossary of key terms from the book is online and can be searched or
browsed.

Online Appendices
Appendix I: A Field Guide to Protein Folds illustrates the structure and topology of commonly
encountered protein folds.
Appendix II: Methods of Structure Determination describes the main methods currently
used in the practice of structural biology organized by computational approaches, electron
microscopy, nuclear magnetic resonance, and X-ray crystallography. Each entry lists key
references for further study.

ACKNOWLEDGMENTS
The authors and publisher of Molecular Biology of Assemblies and Machines specially acknowledge and thank the following people for
their help during the development of the book. Those who provided contributions to chapters are listed first (*), followed by readers (†),
and reviewers (‡).
Chapter 1:
* Neil Rzechorzek (University of Cambridge), ‡ Robert Barber
(University of Wisconsin, Parkside), Jijie Chai (Tsinghua
University), James J. Champoux (University of Washington),
Qintong Li (Sichuan University), Eric May (University of
Connecticut), Jesus Perez-Gil (Complutense University of
Madrid), Jose Rizo-Rey (University of Texas Southwestern
Medical Center), Rajan Sankaranarayanan (CSIR-Centre for
Cellular and Molecular Biology), Sen-Fang Sui (Tsinghua
University), Kunchithapadam Swaminathan (National University
of Singapore), Hongwei Wang (Tsinghua University), Zhisong
Wang (National University of Singapore)
Chapter 2:
* Bradley Cairns (University of Utah School of Medicine), Jeff
Hansen (Colorado State University), Karolin Luger (University
of Colorado, Boulder), Patrick Varga-Weisz (The Babraham
Institute), † Andrew Travers (MRC Laboratory of Molecular
Biology), ‡ Pui Shing Ho (Colorado State University), Hengbin
Wang (University of Alabama, Birmingham)
Chapter 3:
* Stephen D. Bell (Indiana University at Bloomington), David
Jeruzalmi (City University of New York), Anthony Maxwell
(John Innes Centre), ‡ Deepak Bastia (Medical University of
South Carolina), Sue Cotterill (St George’s College, University
of London), Agnieszka Gambus (University of Birmingham),
Stephen Kearsey (University of Oxford), Kenneth N. Kreuzer
(Duke University), Linda Reha-Krantz (University of Alberta),
Phoebe A. Rice (University of Chicago)
Chapter 4:
* Tom Ellenberger (Washington University of St Louis School of
Medicine), Luca Pellegrini (University of Cambridge), Phoebe
A. Rice (University of Chicago), Neil Rzechorzek (University of
Cambridge), ‡ Li Fan (University of California, Riverside), Youri
Pavlov (University of Nebraska Medical Center), John Tainer
(The Scripps Research Institute), Hongwei Wang (Tsinghua
University), Mike Williamson (University of Sheffield), Yanbin
Zhang (University of Miami School of Medicine)
Chapter 5:
* Patrick Cramer (Max Planck Institute for Biophysical Chemistry),
Reinhard Lührmann (Max Planck Institute for Biophysical
Chemistry), Cindy L. Will (Max Planck Institute for Biophysical
Chemistry), † Kiyoshi Nagai (MRC Laboratory of Molecular
Biology), ‡ Robert Barber (University of Wisconsin, Parkside),
Sonja Baumli (University of Oxford), Elena Conti (Max Planck
Institute for Biochemistry), Zhi John Lu (Tsinghua University),
ix
Leticia Márquez-Magaña (San Francisco State University), Daniel
Reines (Emory University School of Medicine), Hongwei Wang
(Tsinghua University), Mike Williamson (University of Sheffield)
Chapter 6:
* Anders Liljas (Lund University), Elizabeth Villa (University
of California, San Diego), † Andreas Bracher (Max Planck
Institute for Biochemistry), Venkatraman Ramakrishnan (MRC
Laboratory of Molecular Biology), Daniel Wilson (Ludwig
Maximilians Universität München), ‡ Scott C. Blanchard (Weill
Cornell Medical College), Jane Dyson (The Scripps Research
Institute), Katrina Forest (University of Wisconsin, Madison), Kurt
Frederick (Ohio State University), Richard Jackson (University
of Cambridge), Daniel Kaganovich (Hebrew University), Harry
Noller (University of California, Santa Cruz), Peter Tompa
(Flanders Institute of Biotechnology)
Chapter 7:
† Gregory Effantin (Institut de Biologie Structurale, Grenoble),
‡ Robert Barber (University of Wisconsin, Parkside), Raymond
Deshaies (California Institute of Technology), Fred Gorelick (Yale
University), Jörg Martin (Max Planck Institute for Developmental
Biology), Mike Williamson (University of Sheffield), Zhaohui Xu
(University of Michigan)
Chapter 8:
‡ Timothy S. Baker (University of California, San Diego), Jay
Brown (University of Virginia), Peter Prevelige (University
of Alabama, Birmingham), Peter Stockley (University of
Leeds), Nicola Stonehouse (University of Leeds), Stephen Stray
(University of Mississippi Medical Center), John A. Tainer (The
Scripps Research Institute)
Chapter 9:
* Edward Bayer (Weizmann Institute of Science, Israel), Raphael
Lamed (Tel Aviv University), Neil Rzechorzek (University of
Cambridge), ‡ Robert Barber (University of Wisconsin, Parkside),
Katrina Forest (University of Wisconsin, Madison), Marco W.
Fraaije (University of Groningen), Perry A. Frey (University of
Wisconsin, Madison), Eric May (University of Connecticut),
Edith Miles (National Institutes of Health), Jesus Perez-Gil
(Complutense University of Madrid)
Chapter 10:
* Martin Beck (European Molecular Biology Laboratory), Elena
Conti (Max Planck Institute of Biochemistry), Oliver Daumke
(Max Delbrück Center for Molecular Medicine), Jenny Hinshaw
(National Institutes of Health), Tomas Kirchhausen (Harvard
Medical School), Jose Rizo-Rey (University of Texas Southwestern
x Acknowledgments
Medical Center), Natalie Strynadka (University of British
Columbia), Thomas Spreter (University of British Columbia),
Elizabeth Villa (University of California, San Diego), † Stuart T.
Endo-Streeter (Duke University), Gabriel Waksman (Institute
of Structural and Molecular Biology, University of London),
‡ Kenneth D. Belanger (Colgate University), Yuh Min Chook
(University of Texas Southwestern Medical Center), Graeme
Henderson (University of Bristol), Eamonn Kelly (University of
Bristol), Robert Kretsinger (University of Virginia), Qingzhen Liu
(Wuhan University), Danton H. O’Day (University of Toronto,
Mississauga), Brian Reilly (Texas Tech University), Karen K.
Resendes (Westminster College, New Wilmington), David
Stephens (University of Bristol), Mike Williamson (University of
Sheffield)
Chapter 11:
* Josephine C. Adams (University of Bristol), Jürgen Engel
(University of Basel), Maria K. Hoellerer (University of Oxford),
Martin E. M. Noble (Newcastle University), Gina E. Sosinsky
(University of California, San Diego), William I. Weis (Stanford
University School of Medicine), † Gabriel Waksman (Institute of
Structural and Molecular Biology, University of London), ‡ Clive
R. Bagshaw (University of Leicester), Iain Campbell (University
of Oxford), Matthias M. Falk (Lehigh University), Peter J. Koch
(University of Colorado, Denver)
Chapter 12:
* David Barford (MRC Laboratory of Molecular Biology),
† Jane Endicott (Newcastle University), John Heath (University
of Birmingham), Carl-Henrik Heldin (Uppsala University),
Brenda Schulman (St Jude Children’s Research Hospital,
Memphis), Stephen Sprang (University of Montana), Susan
Taylor (University of California, San Diego), Nick Tonks (Cold
Spring Harbor Laboratory), ‡ Josephine C. Adams (University
of Bristol), Ye-Guang Chen (Tsinghua University), Christopher
Garcia (Stanford University School of Medicine), Graeme
Henderson (University of Bristol), Eamonn Kelly (University
of Bristol), Robin Ketteler (University College London), John
Kuriyan (University of California, Berkeley), Qingzhen Liu
(Wuhan University), David Stephens (University of Bristol), John
J. G. Tesmer (University of Michigan), Qi Wang (University of
California, Berkeley), Mike Williamson (University of Sheffield)
Chapter 13:
* Anthony A. Hyman (Max Planck Institute of Molecular
Cell Biology and Genetics), Andrea Musacchio (Max Planck
Institute of Molecular Physiology), Laurence Pelletier (Mount
Sinai Hospital), Stefan Riedl (Sanford-Burnham Medical
Research Institute), † Kim Nasmyth (University of Oxford),
‡ Clive R. Bagshaw (University of California, Santa Cruz),
Solomon Bhupanapadu Sunkesula (Brody School of Medicine
at East Carolina University), Robert Friis (University of Bern),
Stephan Geley (Innsbruck Medical University), Leonard R.
Johnson (University of Tennessee College of Medicine), Brett D.
Keiper (Brody School of Medicine at East Carolina University),
Qingzhen Liu (Wuhan University), David Morgan (University
of California, San Francisco), David Stephens (University of
Bristol), Zhaohui Xu (University of Michigan)
Chapter 14:
* Linda A. Amos (MRC Laboratory of Molecular Biology),
Dennis Bray (University of Cambridge), Roger Craig (University
of Massachusetts Medical School), Kenneth C. Holmes (Max
Planck Institute for Medical Research), Dennis R. Thomas (Max
Planck Institute for Biochemistry), ‡ Clive R. Bagshaw (University
of California, Santa Cruz), Jonathon Howard (Yale University),
Hugh Huxley (Brandeis University), Junmin Pan (Tsinghua
University), Mary E. Porter (University of Minnesota, Twin
Cities), Ivan Rayment (University of Wisconsin, Madison), Karl-
Gösta Sundqvist (Karolinska Institute), Ron Vale (University of
California, San Francisco), Zhisong Wang (National University of
Singapore)
Chapter 15:
* Gary Cecchini (The Veteran’s Health Research Institute), Richard
J. Cogdell (University of Glasgow), Ryota Iino (National Institutes
of Natural Sciences, Japan), Hiroyuki Noji (University of Tokyo),
‡ Christoph van Ballmoos (Stockholm University), Giorgio Lenaz
(University of Bologna), Thomas Meier (Max Planck Institute of
Biophysics), Peter Rich (University College London), Alastair
Stewart (Victor Chang Cardiac Research Institute), Zhisong
Wang (National University of Singapore), Joachim Weber (Texas
Tech University)
Chapter 16:
* Konstantinos Beis (Imperial College London), Alex Cameron
(Diamond Light Source), Thomas Sørensen (Diamond Light
Source), Robert M. Stroud (University of California, San
Francisco), Nigel Unwin (MRC Laboratory of Molecular
Biology), † Frances Ashcroft (University of Oxford), ‡ Isaiah T.
Arkin (Hebrew University), Chris Miller (Brandeis University),
Mark Sansom (University of Oxford), G. Graham Shipley (Boston
University), John J. G. Tesmer (University of Michigan), Gareth R.
Tibbs (Columbia University), Giovanni Zifarelli (Italian National
Research Council)
Chapter 17:
* Piet Gros (University of Utrecht), David A. Shore (The Scripps
Research Institute), Robyn L. Stanfield (The Scripps Research
Institute), Ian A. Wilson (The Scripps Research Institute),
† Nick Gay (University of Cambridge), Susan Lea (University of
Oxford), Anton van der Merve (University of Oxford), Bob Sim
(University of Oxford), ‡ Qingzhen Liu (Wuhan University), Mary
Jane Niles (University of San Francisco), Brian Reilly (Texas Tech
University), David Stephens (University of Bristol), Ronald Paul
Taylor (University of Virginia School of Medicine), Zhaohui Xu
(University of Michigan)
Appendix I, A Field Guide to Protein Folds:
* Nick Furnham (London School of Hygiene & Tropical Medicine)
Appendix II, Methods of Structure Determination:
* Bauke W. Dijkstra (University of Groningen), Angela M.
Gronenborn (University of Pittsburgh), Tatyana Polenova
(University of Delaware), Andrej Sali (University of California,
San Francisco)
CONTENTS

Life Processes are Driven by Macromolecular Assemblies and Machines xxvii

Chapter 1 The Machines and Assemblies of Life 1
Chapter 2 Chromatin 49
Chapter 3 DNA Replication 77
Chapter 4 DNA Repair and Recombination 123
Chapter 5 Transcription 173
Chapter 6 Protein Synthesis and Folding 213
Chapter 7 Intracellular Proteolysis: Protein Quality Control and
Regulatory Turnover 263
Chapter 8 Assembly of Viruses 297
Chapter 9 Multienzyme Complexes: Catalytic Nanomachines 357
Chapter 10 Transport 397
Chapter 11 Connectivity and Communication 449
Chapter 12 Signaling 495
Chapter 13 The Cell Cycle and Programmed Cell Death 549
Chapter 14 Motility 597
Chapter 15 Bioenergetics 663
Chapter 16 Membrane Channels and Transporters 721
Chapter 17 Complexes of the Immune System 761
Glossary 799
Index 817
SPECIAL FEATURES

Box 1.1 Isomerism 26

Box 1.2 Metabolic control analysis 34
Box 1.3 Sickle-cell anemia and aberrant polymerization of a mutant hemoglobin 37
Box 1.4 Diffusion: Fick’s laws 38
Box 1.5 Major compartments in eukaryotic cells 41
Box 2.1 The hierarchical condensation of DNA in the eukaryotic nucleus 51
Box 2.2 DNA structure 55
Box 2.3 DNA topology 56
Box 2.4 DNA methylation 72
Box 3.1 ATPases are important components of a wide variety of protein machines 82
Box 5.1 Inhibitors of RNA polymerase 180
Box 5.2 DExD/H box ATPases and helicases 195
Box 5.3 KH, S1, and RRM RNA-binding domains 197
Box 5.4 Alternative splicing and the tau protein 204
Box 7.1 Enzymatic mechanisms of proteolysis 265
Box 7.2 Mechanoenzymes of the AAA+ ATPase family 268
Box 7.3 Proteasome inhibitors as anti-cancer drugs 282
Box 7.4 Endoplasmic reticulum-associated degradation (ERAD) 286
Box 8.1 Assembly and maturation of human immunodeficiency virus (HIV) 344
Box 9.1 Structure and mechanism of E2 in a 2-OADH complex 365
Box 9.2 Structure and mechanism of E1 in a 2-OADH complex 366
Box 9.3 Structure and mechanism of E3 in a 2-OADH complex 368
Box 10.1 The COPI and COPII membrane traffic pathways 401
Box 11.1 The blood–brain barrier 459
Box 12.1 G proteins 500
Box 12.2 Protein kinases 508
Box 12.3 Protein kinase inhibitors for the treatment of cancer 518
Box 14.1 Research on muscle contraction has deep roots 613
Box 15.1 Cofactors of biological oxidation 666
Box 15.2 Shuttle systems exist to transfer the reducing equivalents of NADH into the mitochondrion 669
Box 15.3 Quantum mechanical tunneling by electrons and protons 672
Box 15.4 Singlet and triplet states and intersystem crossing 689
Box 16.1 The flux of ions through a channel when limited by diffusion 722
Box 17.1 Inflammation 762
Box 17.2 Complement nomenclature 769
Box 17.3 Immunoglobulins 781
Box 17.4 A summary of the cells involved in the immune response 784
Detailed Contents

Life Processes are Driven by ‘Single-molecule’ methods interrogate

macromolecules one at a time 23
Macromolecular Assemblies Molecular dynamics models the motions of crystal
and Machines xxvii structures in the presence of a force field 25
1.6 Catalysis 25
Chapter 1 The Machines and
Enzymes form highly specific but transient
Assemblies of Life 1 complexes with their substrates 25
1.1 Expression of the genetic blueprint 1 Enzyme kinetics are governed by a few equations 26
The flow of information is not perfect and not A key feature of enzyme catalysis is the tight
always in one direction 2 binding of the transition state 27
Enzymes generate catalytic rate enhancements
1.2 Weak forces and molecular interactions 3 in multiple ways 28
All weak forces other than hydrophobic interactions Enzymes can be inhibited reversibly and
are electrostatic in origin 4 irreversibly 28
Hydrophobic interactions drive the folding and Coupling of enzyme-catalyzed reactions allows
assembly of macromolecules 5 energetically unfavorable reactions to occur 30
The energy balance in folding and assembly has both
enthalpic and entropic contributions 7 1.7 Signaling and regulatory mechanisms 30
Size and topography matter for interaction patches 7 Ligand-induced conformational change and
cooperativity are widespread methods of controlling
A certain minimum strength of interaction is required biological activity 30
for specificity 8
Allosteric proteins are regulated by a special form of
Cooperativity enhances stability in multi-subunit cooperativity 31
complexes 9
Allosteric enzymes do not follow Michaelis–Menten
1.3 Protein folding and stability 10 kinetics 32
Protein folding follows pathways populated with Allostery is mediated by protein/protein interactions
intermediates 10 and conformational changes 32
Protein structures are only marginally stable 12 Reversible covalent modification controls the
activities of some proteins 34
Protein stability correlates with size and other
factors such as covalent cross-links 12 Homeostasis is an important aspect of response to
environmental change 34
Many cellular proteins denature collectively under
thermal stress 13 1.8 Macromolecular crowding 35
Proteins from thermophilic organisms are not very Molecular crowding affects reaction rates, protein
different from mesophilic homologs 14 folding, assembly, and stability 36
1.4 Self-assembly and symmetry 14 Macromolecular crowding affects diffusion rates 36
Most proteins form symmetrical oligomers with Models of crowded intracellular environments can
two or more subunits 14 now be built 39
Symmetry defines a set of larger structures composed 1.9 Cellular compartmentation and
of multiple copies of identical subunits 15 evolution 39
Line and cyclic point group symmetries generate All cells belong to one of the three Urkingdoms:
helices and rings 15 Archaea, Bacteria, Eukarya 40
Cubic symmetry is employed in a variety of Bacteria have an open compartment, the nucleoid,
oligomeric proteins 17 and a membrane-delimited compartment, the
Assembly proceeds along pathways 18 periplasm 40
Why are there so many large macromolecular Archaea more closely resemble eukaryotes than
assemblies? 18 bacteria in some key features 42
Major differences exist between archaeal and
1.5 Macromolecular dynamics 19 bacterial cell envelopes 43
Ensemble methods measure the net signal from Eukaryotic cell organelles probably arose by engulfing
numerous contributors 21 bacteria 44
xiv Detailed Contents

Higher eukaryotes have similar numbers of genes 2.4 Epigenetic mechanisms 68

as lower eukaryotes but many more regulatory Histone post-translational modifications are carriers
elements 45 of epigenetic information 69
References 46 Special protein domains recognize specifically
modified histone residues 69
Chapter 2 Chromatin 49 The histone code hypothesis suggests that the PTM
pattern of a nucleosome acts as a ‘barcode’ 70
2.1 Introduction 49 Methylation of CpG islands is another epigenetic
marker that results in widespread gene silencing 71
2.2 Nucleosomes and higher order
chromatin structures 50 X-chromosome inactivation and imprinting are
important epigenetic phenomena in mammalian cells 73
The core histones have a two-domain organization
and structure 50 2.5 Summary 74
Core histones assemble into H2A–H2B and H3–H4 References 75
heterodimers and form a metastable histone octamer 52
A nucleosome is 147 bp of DNA wrapped around a
histone octamer 53 Chapter 3 DNA Replication 77
Nucleosomal DNA is a highly distorted superhelix 53 3.1 Introduction 77
Nucleosomes are assembled sequentially with the
help of histone chaperones 54 3.2 Initiation and elongation 78
The structure of the nucleosome is intrinsically A series of multiprotein complexes are recruited to
dynamic 57 the bacterial origin of DNA replication 78
DNA sequence directs specific positioning of The active form of DnaA is an ATP-dependent
nucleosomes in vitro and in vivo 58 oligomer 79
Nucleosomal arrays form higher order structures that DNA replication in eukaryotes proceeds from
differ in their degree of condensation 58 multiple origins 80
Linker histones stabilize condensed 30 nm chromatin The origin recognition complex is the homolog of the
structures 58 DnaA oligomer in eukaryotes 81
The structure of the 30 nm fiber remains unsettled 59 Bacterial DnaA melts DNA at the origin to enable
The 30 nm fiber has a heteromorphic structure loading of the DnaB replicative helicase 85
dependent on nucleosome repeat length and packing Eukaryotic DNA is licensed for replication when an
order 59 inactive replicative helicase is loaded at the ORC in
Higher order folding of nucleosomal arrays is the G1 phase of the cell cycle 86
regulated by cations and the core histone N-terminal Helicases use the energy of nucleotide binding and
tail domains 60 hydrolysis to unwind duplex DNA 87
Core histone isoforms have variant sequences and The MCM helicase is activated in S phase of the cell
functions 61 cycle 88
Core histones undergo many specific post-translational The replicative helicases of eukaryotes and archaea
modifications with structural implications 62 track on DNA in a 3′ to 5′ direction 89
Chromatin architectural proteins are essential for Papillomavirus E1 helicase tracks on ssDNA in the
higher order chromatin structures 62 3′ to 5′ direction 89
Genomic chromatin is a heterogeneous and complex Single-stranded DNA is bound by a protective protein
macromolecular assembly 63 before it enters the DNA polymerase 90
Elucidating chromosomal architecture beyond the The eukaryotic ssDNA-binding protein RPA also
30 nm fiber remains a challenge 63 helps organize many other proteins in the
2.3 Remodeling complexes 64 replication fork 91
Remodelers regulate DNA exposure in chromatin 64 The RNA primers for DNA polymerases are
synthesized by primase, a special polymerase 92
Remodelers can be separated into four families defined
by their composition and activities 65 Primases make primers of defined length but exhibit
low fidelity in the copying process 92
Remodelers have specialized as well as common
properties 66 DNA is copied by DNA polymerases 93
The disruption of histone–DNA contacts is DNA polymerases contain a 3′ to 5′ exonuclease as
ATP-dependent 66 well as a 5′ to 3′ polymerase activity 94
Remodeler regulation depends on the interplay with The high fidelity of replicative polymerases arises from
histone post-translational modifications 67 multiple sources 95
Structural models inform how remodelers engage and DNA replication is continuous on one strand but not
remodel nucleosomes 68 the other 97
 Detailed Contents xv

Numerous enzymes interact at a replication fork and Chapter 4 DNA Repair and
function as a concerted giant assembly 97
DNA is synthesized simultaneously on leading and
Recombination 123
lagging strands 98 4.1 Introduction 123
Three different DNA polymerases are required for Lesions can occur in one or both strands of duplex
DNA replication and operate differently on the DNA and are repaired by five different enzyme
leading and lagging strands 99 systems 123
The processivity of DNA polymerases is enhanced by
a sliding clamp 99 4.2 Direct reversal of damage in one
strand of duplex DNA 126
DNA Pol ε and Pol δ, together with many other
proteins, associate with a sliding clamp on the DNA 100 Lesions induced by UV light can be repaired directly 126
The β clamp and DNA polymerase are loaded onto Some aberrant methylations can also be repaired
the RNA-primed DNA by an ATP-dependent clamp directly 127
loader 101 4.3 Templated repair of lesions affecting
The clamp loader forms an ATP-dependent spiral one strand of the DNA duplex 130
structure round the primer–template junction 103 Base-excision repair is initiated by DNA glycosylases 130
The clamp loader is bound to SSB by a heterodimer DNA glycosylases search for lesions in DNA by
of χ and ψ proteins 105 transient encounters, both passive and active 131
DNA Pol I and DNA ligase are required to fill in and Replacement of the excised base requires additional
close the gaps between Okazaki fragments on the enzymes 131
lagging strand 105
Nucleotide-excision repair deals with bulky lesions 133
The replisome is held together by an array of
protein/protein interactions 107 UvrABC interact and function sequentially in
bacterial NER 133
A specialized set of proteins is required for RNA
Many proteins act sequentially in eukaryotic NER 135
primer excision 107
Interaction of damaged DNA with XPC (Rad4) is
Eukaryotic DNA ligases resemble E. coli LigA but
the first step in eukaryotic NER 135
use ATP rather than NAD+ as co-substrate 108
Mismatch repair corrects mispaired bases that are left
3.3 Termination of DNA replication 109 uncorrected during DNA replication 136
Bacterial chromosomes contain termination sites for MutS forms an asymmetric homodimer and
DNA replication 109 mismatch recognition involves interaction with only
The linear DNA in eukaryotic chromosomes is one subunit 138
replicated by a special mechanism that also protects 4.4 Repair of double-strand breaks 139
its ends 110
Recognition of the broken DNA ends is the first step
Telomeric DNA is synthesized and maintained by in DSB repair 139
telomerase, a specialized polymerase that utilizes RNA
as a template 111 The Mre11 component of the MRN complex is a
dimeric nuclease that can bind DNA ends 141
Replicons and factories 112
Rad50 is an ATPase that interacts with Mre11 and
3.4 DNA topology in replication 113 undergoes reversible ATP-dependent dimerization 142
DNA topoisomerases overcome topological The Nbs1 component helps link the MRN complex
constraints in DNA replication 114 to the appropriate DSB response 143
DNA topoisomerases fall into two classes with The Ku protein mediates DSB repair by the NHEJ
similar but different mechanisms 114 pathway 144
Type IA topoisomerases cleave one strand of the DNA-bound Ku recruits other factors essential
duplex and pass the other strand through the gap for NHEJ 144
created before religation 114 MMEJ is a variant of NHEJ 145
Type IB topoisomerases cleave one strand of the HR operates by DNA strand exchange in all three
duplex and allow part of the duplex DNA to undergo kingdoms of life 146
controlled rotation before religation 116
DNA strand exchange is catalyzed by filaments of
Type II topoisomerases cut both strands of dsDNA a RecA family ATPase 146
and pass one segment of DNA through the gap
created 117 A synaptic complex between a RecA nucleoprotein
filament and sister chromatid DNA promotes
The C-terminal domains of bacterial topoisomerase fidelity in strand exchange 147
IIA enzymes impose their specific biological
functions 119 Coordinated activities of helicases and nucleases
generate the long 3′ overhangs for strand invasion 149
3.5 Summary 121 Assembly and disassembly of the recombinase
References 122 nucleoprotein filament are tightly regulated 150
xvi Detailed Contents

Four-way (Holliday) junctions are key intermediates Inhibitors of RNA polymerase have helped to define
in DSB repair and meiosis 151 mechanisms 178
Branch migration is driven by the RuvAB complex 151 The polymerase can overcome natural obstacles to
HJs are resolved by specific endonucleases or transcription elongation 180
dissolved by a helicase/topoisomerase 153 RNA synthesis has a higher error rate than DNA
Homology repair also restarts stalled replication synthesis 181
forks 154 Small RNAs can inhibit transcription 182
Damage tolerance and error-prone translesion DNA Messenger RNA is protected at the 5′ end by a cap
synthesis 155 structure 182
4.5 Site-specific DNA recombination Termination is closely associated with 3′-
and DNA transposition 156 polyadenylation of pre-mRNA 182
Tyrosine recombinases cut one strand of each Pol I and Pol III are similar in structure to Pol II but
partner duplex at a time 157 synthesize different RNAs 183
Formation and resolution of the HJ DNA 5.3 The pre-initiation complex 184
intermediate requires little movement in the The prokaryotic initiation complex involves just the
recombinase tetramer 159 σ factor and polymerase 184
Bacteriophage λ integrase makes use of ancillary The eukaryotic PIC includes many different
DNA-bending proteins 160 transcription factors 185
Unusual tyrosine recombinases act as resolvases of TFIID acts as a scaffold for the assembly of other
DNA replicon dimers 161 general transcription factors 186
Integron integrases are tyrosine recombinases that TFIIA and TFIIB help stabilize the TBP/TATA
recognize hairpinned ssDNA substrates 162 complex 187
Hairpin telomeres at the end of linear TFIIB recruits the promoter complex to Pol II 187
chromosomes are resolved by specialized tyrosine
recombinases 162 TFIIH contains enzymes that unwind DNA and
phosphorylate the Pol II CTD 188
Serine recombinases overlap tyrosine recombinases
in biological function but differ evolutionarily and Transcription elongation requires protein kinase
mechanistically 163 activity of Cdk9/cyclin T (P-TEFb) 189
Gene-specific transcription factors regulate
Serine recombinases can be regulated by means of
transcription 189
accessory proteins 165
The Mediator complex links gene-specific
Accessory proteins can direct a serine recombinase
transcription factors to the PIC 191
to catalyze inversion rather than deletion of a DNA
segment 166 5.4 RNA processing: the spliceosome 192
Another family of transposases and retroviral The pre-mRNA introns are removed in a two-step
integrases is defined by a DDE motif in the process 192
active site 167
Assembly and disassembly of the spliceosome
DDE transposases and retroviral integrases are proceed in stepwise fashion 192
diverse but share some structural features 168
A dynamic spliceosomal RNA/RNA interaction
Mobile elements are responsible for a large network is formed during splicing 195
proportion of important evolutionary changes in
Splice-site recognition in the E and A complexes
genomes 170
involves the coordinated action of RNA and protein 196
4.6 Summary 170 The spliceosome has a complex and dynamic protein
References 171 composition 198
A two-state model for the catalytic center of the
spliceosome 198
Chapter 5 Transcription 173 The spliceosome appears to act mostly as a ribozyme 199
5.1 Introduction 173 A crystal structure of the U1 snRNP suggests a
mechanism for 5′ss recognition 200
5.2 RNA polymerase II (Pol II) and the Spliceosome assemblies have been visualized by
elongation complex 174 electron microscopy and labeling experiments 202
RNA polymerases are multi-subunit enzymes that
Splicing enhancers and silencers regulate alternative
share a conserved core 174
splicing 203
The elongation complex binds template DNA,
nucleotide triphosphates, and newly synthesized RNA 176 5.5 The exosome 204
Nucleotide selection is coupled to catalysis 177 Exosomes are based on a hexameric ring structure 204
Nucleotide addition and translocation require a The exosome has processive 3′ → 5′ exoribonuclease
dynamic polymerase catalytic site 177 activity 205
 Detailed Contents xvii

The archaeal exosome uses RNA-binding proteins Some proteins require the assistance of chaperonins
to target RNA to the catalytic site 206 to reach their native states 235
There are similarities in mechanisms of the exosome Chaperonins are evolutionarily conserved
and the proteasome 208 protein-folding machines 235
The human exosome core is similar in structure to GroEL is an ATP-driven folding machine 236
the bacterial and archaeal exosomes but has no The thermosome is the archetype of group II
phosphorolytic catalytic activity 208 chaperonins 237
Additional subunits provide nuclease hydrolytic Group II chaperonins undergo large-scale
activity 209 conformational changes 238
5.6 Summary 210 The chaperonin TRiC/CCT of eukaryotes is built
References 211 from eight distinct subunits 239
Proteins with complex fold topologies are dominant
clients of TRiC/CCT 240
Chapter 6 Protein Synthesis Hsp90 chaperones regulate the activity of
and Folding 213 multifarious client proteins 240
Hsp90 undergoes large-scale conformational changes
6.1 Introduction 213 during its reaction cycle with the help of its
co-chaperones 241
6.2 The ribosome 214
Small heat shock proteins protect client proteins
tRNAs are adaptor molecules between genes and from aggregation 242
proteins 214
α-Crystallin domains form the core of all sHsps 243
tRNA synthetases charge tRNAs with their cognate
amino acids 215 6.4 Natively unfolded proteins 243
The ribosome is an ancient molecular machine that Natively unfolded regions may be recognized by
catalyzes protein synthesis 215 multiple experimental approaches 243
X-ray crystallography and electron microscopy Natively unfolded proteins have distinctive amino
have revealed the structure of ribosomes 217 acid compositions 244
The ribosome is a ribozyme 220 Natively unfolded regions are often involved in
Translation factors enhance the efficiency of protein regulation, folding when they engage interaction
synthesis 221 partners 244
Translation occurs in four stages: initiation, 6.5 Protein misfolding and amyloid fibrils 247
elongation, termination, and recycling 222 Amyloids share distinctive properties: fibril
Initiation factors bind to the small subunit 222 morphology, stability, dye-binding, and cross-β
The subunits associate to form the 70S ribosome conformation 248
during initiation 223 Amyloid fibrils are polymorphic 250
Elongation factors escort tRNAs into and within Models of amyloid fibrils envisage differing
the ribosome 224 configurations of β strands 250
Amino acids are added to the nascent protein The native folds of β-solenoid proteins are amyloid-like 252
during elongation 224 Fibril assembly proceeds in two phases: nucleation
tRNA needs to deform to be probed for codon– (slow) and elongation (a faster, templated process) 253
anticodon complementarity 225 Oligomeric assemblies may be the pathogenic
The ribosome decodes the signal 225 agents in some neurodegenerative and other
amyloid-related diseases 254
Peptide bond formation occurs rapidly and
spontaneously 226 For a growing number of proteins, amyloid
represents the native functional state 254
The ribosome rocks during translocation 226
Release and recycling factors recognize and execute 6.6 Prions 255
the end of the cycle 227 Prion domains are unfolded in the wild-type
A protein is born and the cycle begins again 228 protein and amyloid in the prion 256
Proteins can be translocated into and across The prion domains self-assemble to form amyloid
membranes 229 fibril backbones of prion filaments 257
Multiple ribosomes assembled into polysomes Prion infection is accompanied by a loss or gain of
translate the same mRNA simultaneously 230 function 259
Many antibiotics target the ribosome and inhibit In infection, fibrillation is nucleated by transmitted
its function 230 or spontaneously formed seeds 260
How widespread are prions? 260
6.3 Molecular chaperones 232
Nascent polypeptide chains are met by an array of 6.7 Summary 260
chaperones 234 References 261
xviii Detailed Contents

Chapter 7 Intracellular Proteolysis: Chapter 8 Assembly of Viruses 297

Protein Quality Control and 8.1 Introduction 297
Regulatory Turnover 263 8.2 Principles of virus replication 298
7.1 Introduction 263 Viruses behave like machines and self-replicating
automata 299
7.2 Principles of unfoldase-assisted
proteolysis 264 Helical and icosahedral symmetry are widely
employed in virus architecture 300
Classical proteases selectively sever peptide bonds 264
Spherical capsids are protein shells with icosahedral
Substrate specificity is conferred by the regulatory symmetry 301
particles 264 Quasi-equivalence allows the assembly of larger
Unfoldase-assisted proteases have stacked-ring capsids with more than 60 subunits 303
architecture and modular organization 266 Larger viruses show progressively greater complexity
7.3 Unfoldase-assisted proteases in in structure and composition 305
bacteria and eukaryotic organelles 267 Pairs of complementary interaction patches are the
Assembly involves polymorphisms and symmetry key to self-assembly 305
mismatches 270 Pathways are mapped by characterizing mutants for
Substrate proteins are marked for degradation by which assembly is blocked 306
peptide signals 270 Many capsids are initially assembled as precursor
Accessory domains and adaptor proteins affect procapsids that subsequently mature 307
substrate selection 272 Capsids and crystals exhibit defects, symmetry-
Substrates are unfolded and translocated along an breaking, and dynamics 307
axial pathway 273 8.3 Helical viruses 308
Unfoldase-assisted proteases are machines with TMV is a rigid rod containing a single-stranded
moving parts 274 RNA and only one type of capsid protein 308
Proteolytic active sites are sequestered inside gated Polarity of the helical array is important for
chambers 275 disassembly in vitro and in vivo 309
Proteases have regulatory roles in the replication TMV RNA is selected for encapsidation by
cycles of bacteria and bacteriophages 277 recognition of an internal stem-loop structure 310
7.4 The proteasome 278 Filamentous bacteriophages have long flexuous
capsids enclosing circular ssDNA genomes 311
The molecular architecture of 20S proteasomes is
conserved from archaea to humans 278 Attachment of filamentous bacteriophages to pili
initiates infection 312
α subunits and β subunits have the same basic fold 279
Encapsidation of the DNA proceeds via a
The N-terminal threonine functions as a nucleoprotein filament intermediary 314
single-residue active site 280
Ff virions assemble in the inner membrane and are
Assembly of the complex precedes active site formation 281 secreted through the outer membrane 316
Access to the proteolytic chamber is controlled by Packaging of ssDNA in Ff bacteriophages involves
gated pores 283 charge-matching 317
The 11S regulator acts as a gate opener 283 The ssDNA in bacteriophages Pf1 and Pf3 may be
PAN is an archetypal proteasome-activating inside-out 318
nucleotidase 284 8.4 Small icosahedral viruses 318
The 19S regulatory particle links the ubiquitin Capsid proteins may self-assemble or co-assemble
system with the proteasome 285 with the genome 318
The base subcomplex recruits substrates and prepares Nodaviruses have ordered RNA and mature by
them for degradation 285 autocatalytic proteolysis 321
The lid subcomplex serves to deubiquitylate substrates 288 The capsid protein of a simple plant virus has
7.5 Giant proteases 289 polymorphic assembly products 322
Hepatitis B virus capsid is a porous compartment for
In archaea, tricorn protease is the archetypal giant
retrotranscription 323
protease 289
Picornavirus assembly involves proteolytic
Tripeptidyl peptidase II is an enzyme that counts in processing of a polyprotein 324
threes 291
Breathing capsids fluctuate in their ground-state
TPPII has an unusual spindle-like architecture 291 conformations 326
TPPII activity increases with assembly 293
8.5 Large icosahedral viruses 326
7.6 Summary 293 Assembly proceeds in three stages: procapsid
References 294 assembly, DNA packaging, and maturation 327
 Detailed Contents xix

Procapsid assembly involves portal, capsid protein, Substrate channeling takes place through molecular
scaffolding protein, and protease 328 tunnels or by the covalent attachment of
Procapsid assembly is assisted by host chaperones intermediates to swinging arms 358
and scaffolding proteins 329 9.3 Multienzyme complexes with tunnels 360
Capsid maturation involves a massive conformational Tryptophan synthase has two active sites connected
change 330 by a molecular tunnel 360
Herpesvirus capsids assemble in the cell nucleus The tunnel in tryptophan synthase is gated 361
along a phage-like pathway 331
Ammonia is channeled as a reaction intermediate
Some viral DNAs are packed in coaxial spools by in several different enzyme complexes 361
terminase, a motor protein 333
The capsid architecture of adenovirus is shared by 9.4 Multienzyme complexes with lipoic
some dsDNA bacteriophages 333 acid or biotin in their swinging arms 363
Lipoic acid-dependent 2-oxo acid dehydrogenase
8.6 Double-stranded RNA viruses 334 multienzyme complexes are built round
Double-stranded RNA viruses have one, two, or multisubunit cores 363
three nested protein shells 336 Substrate channeling depends on mobility of the
Conformational changes and proteolytic processing lipoyl domain 363
promote infectivity 337 Multiple active sites are coupled irrespective of their
dsRNA virus capsids are protein-bound geometric arrangement 369
compartments for transcription and replication 338 The intact PDH complex is built of spherical protein
Conformational changes of the procapsid may shells 370
regulate RNA selection by phage ϕ6 339 Eukaryotic PDH complexes have additional
Actively transcribing and replicating viral particles components and are subject to regulation by
are highly dynamic processes 340 reversible phosphorylation 371
8.7 Enveloped viruses 341 Some 2-OADH complexes are based on octahedral
rather than icosahedral symmetry 371
Envelopes are essential for the transport of
nucleocapsids into and out of cells 342 A lipoylated protein is part of a glycine decarboxylase
system in serine biosynthesis 372
Envelope glycoproteins recognize hosts and fuse
membranes 343 Biotin-dependent carboxylases also have a swinging
arm mechanism 373
Influenza virus hemagglutinin, a fusogen, undergoes
pH-dependent conformational changes 346 Allosteric regulation of pyruvate carboxylase involves
structural rearrangements 374
Class II glycoproteins are arranged in icosahedral
lattices 348 Swinging arms extend from flexibly tethered lipoyl
and biotinyl domains to enter active sites 375
8.8 Virus engineering and nanotechnology 349
9.5 Multienzyme complexes with
Phage capsid proteins can be engineered to display phosphopantetheine swinging arms 376
peptides and proteins 349
Fatty acid synthases come in two different forms 376
Full-length proteins can be displayed on icosahedral
capsids 350 Animal FAS is a dimer of multifunctional
polypeptide chains 377
Virus-like particles can be used to generate
protective vaccines 352 Fungal FAS is an α6β6 double-domed cage 378
Virus-like particles are also used in gene therapy and Flexible tethering of the ACP is an essential feature
have applications in drug delivery and clinical of the catalytic mechanism of FAS 380
imaging, and as nano-technological devices 353 The ACP can sequester the long acyl group on the
For VLPs to be used in humans, certain criteria must phosphopantetheine arm and present it for reaction 381
be met 353 Acyl chain length is important in product release 381
Fatty acid degradation partly resembles fatty acid
8.9 Summary 354 synthesis 383
References 355 Substrate channeling in the FAO complex is slightly
leaky 384
Chapter 9 Multienzyme Complexes: Polyketide synthases are related to animal FAS 385
Non-ribosomal peptide synthases also have
Catalytic Nanomachines 357 phosphopantetheine swinging arms 387
9.1 Introduction 357 9.6 The cellulosome 388
9.2 Active-site coupling and substrate The cellulosome is a multienzyme complex
channeling in protein complexes 357 assembled on an inactive protein core 388
Multienzyme complexes channel substrates and Different bacteria generate different sorts of
protect labile intermediates 358 cellulosomes capable of extensive heterogeneity 390
xx Detailed Contents

The type I cohesin/dockerin interaction is plastic SNAREs are recycled by NSF and SNAPs 418
and not confined to cellulosomes 391 Munc18 orchestrates SNARE complex assembly
The carbohydrate-binding module (CBM) anchors together with Munc13 419
the cellulosome to the polymeric substrate 392 Rab is involved in the docking of synaptic vesicles at
Cellulosomes possess a wide range of cellulolytic some presynaptic active zones 420
activities 392 Munc13 and RIM govern synaptic vesicle priming
The modular construction of multienzyme and presynaptic plasticity 421
complexes opens the way to their redesign for Synaptotagmin triggers Ca2+-dependent
selected purposes 393 neurotransmitter release 422
9.7 Summary 393 Complexin plays both active and inhibitory roles 423
References 394 How did synaptic vesicle fusion arise? 424
10.5 Nuclear pore complexes 424
Chapter 10 Transport 397 The nucleus is the defining feature of eukaryotic cells 424
Nuclear pore complexes are the gateways for
10.1 Introduction 397 nucleocytoplasmic transport 425
10.2 Clathrin-mediated endocytosis 398 The NPC is a large and elaborate transport machine 425
Clathrin-coated pits and vesicles are transient The many Nups are built up from domains with only
molecular assemblies that transport a wide range of a few folds 427
different cargoes 398 Nups form stable subcomplexes 429
The building blocks of clathrin-coated structures are NPCs have a modular architecture and are
‘triskelions’ 398 organized dynamically 430
The main motif of the clathrin heavy chains is an 10.6 Nuclear import and export 432
extended α-helical zigzag 399 Nucleocytoplasmic transport is mediated by
Assembly of clathrin-coated structures and transport factors 432
recruitment of cargo requires helper proteins 400 A Ran GTP gradient acts as a cargo pump 432
Heterotetrameric adaptors are the most abundant Importins interact with cargo directly or via adaptor
non-clathrin components of coated vesicles and molecules 433
mediate interaction with endocytic signals 403
Importins adopt shapes complementary to their
Reconstituted coated vesicles provide insights into cargoes 434
interactions in a clathrin lattice 404
The binding of cargo and RanGTP to importins is
Clathrin coats isolated from cells vary greatly in mutually exclusive 434
size and shape 405
The mode of RanGTP binding is similar amongst
Clathrin boxes mediate interactions with heavy chains 406 karyopherins 435
Auxilin and Hsc70 are required for uncoating 407 Cse1 is required for the export of importin α 435
Membrane invagination and budding have Xpot and Xpo5 export tRNAs and pre-micro RNAs 436
substantial energy costs 407
Crm1 exports nuclear proteins that carry an export
10.3 Dynamins are versatile molecular signal 437
machines 408 The karyopherins bind to the FG repeats of the
Dynamin and dynamin-like proteins (DLPs) share nuclear pore complex 437
structural and mechanistic features 409
10.7 Bacterial export and secretion
GTP binding and hydrolysis drive the constriction complexes 438
of the dynamin polymer 410
The Sec system is a general purpose secretion system
Structure and mechanism of other DLPs 412 in bacteria 438
Bacterial dynamin-like protein undergoes The twin-arginine system translocates folded proteins
alternative conformational changes when bound across bacterial inner membranes 439
to lipid membranes 413
The type 3 secretion system (T3SS) is a bacterial
10.4 The machinery of synaptic vesicle nanoinjector 439
fusion 414 The T3SS-associated ATPase is needed for the
Neurotransmitter release is an exquisitely regulated recruitment of secretion substrates 440
form of membrane fusion 414 The basal complex of T3SS is a stack of variably
The release machinery includes a conserved core sized rings with a modular architecture 440
and components specialized for its tight regulation 415 Insertion of the T3SS translocon into the host cell
Membrane fusion is believed to occur through a membrane is mediated by the needle and an adaptor
stalk mechanism 416 at its tip 442
SNAREs are central components of the membrane The type 4 secretion system transports DNA and/or
fusion apparatus 416 proteins across the cell envelope 443
 Detailed Contents xxi

The outer membrane core complex of T4SSs spans 11.5 Gap junctions 466
both membranes of Gram-negative bacteria but Gap junction channels are composed of paired
primarily locates in and near the outer membrane 443 hexameric rings (connexons) 466
The inner membrane complex of T4SS is composed of Gap junctions allow a variety of molecules to pass
VirB3, VirB4, VirB6, VirB8, and the VirB10 N termini 444 between communicating cells 467
The translocation pathway of T4SS switches between Gap junction channels were one of the first
a pilus biogenesis and a DNA transfer mode 444 membrane protein complexes to be characterized by
The type 5 secretion system (T5SS) is a simple but electron microscopy and X-ray diffraction 468
diverse family of transporters 445 Connexon conformations are influenced by changes
Multi-subunit complexes couple the biosynthesis in [Ca2+], pH, phosphorylation state, and binding
and the export of capsular polysaccharides 446 partner 469
10.8 SUMMARY 446 The connexin-like superfamily includes innexins,
pannexins, and vinnexins 470
References 447
11.6 Focal adhesions 471
Focal adhesions are large assemblies with complex
Chapter 11 Connectivity and protein compositions 471
Communication 449 Focal adhesion assembly starts with integrin
activation and is orchestrated by adaptor proteins 472
11.1 Introduction 449
Binding of FAs to the actin cytoskeleton and actin
The size and shape of a eukaryotic cell are determined polymerization are coupled processes 474
primarily by its cytoskeleton 449
The binding sites of FA adaptors are exposed in
Eukaryotic cells communicate with each other at ‘active’ conformations and sequestered in ‘inactive’
specialized contact regions 450 conformations 475
Cells secrete macromolecules that allow them to Small GTPases coordinate the assembly of the actin
communicate with other cells and sense their cytoskeleton and FAs 476
environment 450
Protein phosphorylation has a central role in FA
11.2 Intermediate filaments 451 regulation and signaling 477
IF proteins have a conserved coiled-coil rod domain, Focal adhesions are signaling centers 477
flanked by highly variable N-terminal heads and
C-terminal tails 452 11.7 The extracellular matrix 478
Most IF proteins, except lamins, assemble into ECM is built from a diverse assortment of fibrous
proteins, glycoproteins, and proteoglycans 478
nonpolar filaments 453
Collagens all have triple-helical domains but
IFs have backbones of packed rod domains
assemble into diverse higher-order structures 479
surrounded by protruding end domains 454
Fibrillins constitute a major fibrillar system in
IFs organize into higher-order structures that many connective tissues 481
integrate cells and tissues 455
Fibronectin evolved later than other ECM
Mutations in IF genes underlie numerous human proteins and is specifically a component of
diseases 455 vertebrate ECM 482
11.3 Tight junctions 456 Basement membranes, a specialized ECM, support
Tight junctions consist of networks of paired epithelial and endothelial cell layers 483
intramembrane strands 456 ECM assemblies are remodeled through the actions
Junctional adhesion molecules (JAMs) serve as virus of extracellular proteases 483
receptors 458 Integrins are bidirectional signaling machines 485
In invertebrates, tight junctions are replaced by In mechanotransduction, physical stimuli elicit
septate junctions 460 biological responses such as gene expression 485
11.4 Adherens junctions and 11.8 Bacterial pili 486
desmosomes 461 The binding specificity of pili is conferred by
Classical cadherins and desmosomal cadherins have adhesin proteins 487
similar multi-domain structures 462 Assembly of P-pili and type 1 pili follows the
Cadherin-mediated cell–cell adhesion is homophilic ‘chaperone/usher’ secretory pathway 488
and Ca2+-dependent 463 Pilin subunits are incorporated at the cell-proximal
Cadherins in junctions have both trans and cis end of the growing pilus by a ‘donor strand
interactions 463 exchange’ mechanism 488
Cadherins interact with the actin cytoskeleton via Type 4 pilins have α-helical N-terminal extensions
catenins and other proteins 464 that pack to form the pilus backbone 489
Desmosomes are coupled to the intermediate Phase variation allows the antigenic character of pili
filament network 465 to change without altering their basic architecture 489
xxii Detailed Contents

Pilus retraction by motor-driven depolymerization The βc subunit is common to another set of cytokine
is the mechanism for twitching motility 490 receptor assemblies 530
11.9 Summary 491 Signaling through JAKs and STATs induces
transcription of target genes 531
References 492
TGFβ signals through a combination of RI and
RII Ser/Thr kinase receptors 533
Chapter 12 Signaling 495 Phosphorylation of SMADs by RI induces
transcription of target genes 534
12.1 Introduction 495
DNA recognition by SMADs is highly unusual 535
12.2 Signaling through G-protein-coupled
receptors 496 12.5 Ubiquitylation 536
Light absorption by its retinal cofactor induces a Protein ubiquitylation is catalyzed by a series of
conformational change in rhodopsin 496 three enzymes 537
The G-protein-coupled adrenergic receptors are Activation of UBLs depends on a major
sensitive to agonists and antagonists 498 conformational change in E1 538
Binding of an agonist to the β2-adrenergic receptor In E3 cullin RING ligases, a central cullin subunit
creates a G protein-binding site 498 links the substrate recognition complex and the
Heterotrimeric G proteins act as molecular E2-binding RING domain 538
transducers that couple the activation of GPCRs to CRLs are assembled on a rigid cullin subunit and can
intracellular responses 500 be prevented from assembling by an inhibitor protein 540
The Gα subunit adopts different conformations in Some F-box subunits use WD40 domains to
the GDP- and GTP-bound complexes, whereas Gβγ recognize phosphoprotein substrates 541
is essentially unchanged 503 Neddylation activates CRLs by releasing the RING
Signaling from G proteins to adenylyl cyclase domain of Rbx1 542
produces the second messenger, cyclic AMP 504
The APC/C is a large multi-subunit CRL regulated
Protein kinase A is activated by cAMP and triggers a by phosphorylation and co-activators 543
protein kinase cascade 506
The APC/C is assembled from subunits containing
A protein kinase A-anchoring protein coordinates multiple sequence repeats and undergoes major
multiple effector proteins, and phosphatases and structural change 544
phosphodiesterases terminate the signal 510
PKA activates phosphorylase kinase, which then 12.6 Summary 546
activates glycogen phosphorylase 511 References 547
Glycogen phosphorylase undergoes conformational
changes in response to phosphorylation and
allosteric effectors 512 Chapter 13 The Cell Cycle and
12.3 Signaling through tyrosine kinases 514 Programmed Cell Death 549
Receptor tyrosine kinases are activated by 13.1 Introduction 549
dimerization 514
The insulin receptor tyrosine kinase domain (IRK) is 13.2 The cell cycle 549
a model for tyrosine kinase activation 515 13.3 Transient assemblies of kinases,
The EGFR kinase is activated through asymmetric cyclins, and phosphatases 551
dimerization 516 Mitogenic signals initiate the cell cycle 553
Src is regulated by domain interactions and Cyclin destruction inactivates CDKs 553
phosphorylation 519
Cdk2/cyclin A is a model for the structural basis
Growth factor receptor tyrosine kinases signal to the of CDK regulation 554
small G protein Ras 520
Cyclins can bind kinase substrates at a remote
The Ras/RAF/MEK1/ERK2 signaling pathway leads hydrophobic patch 555
to activation of transcription 522
Activation of MEK and ERK2 leads to activation of Cell cycle inhibitors regulate CDK activity 556
transcription 523 Phosphorylation and dephosphorylation regulate
Growth factor receptor responses also activate at CDK activity 557
least two other pathways 524 Cdk1/cyclin B is the master kinase in mitosis but
other kinases play critical roles at other stages 558
12.4 Cytokine signaling 525
Phosphatases dephosphorylate substrates to
Class I and class II cytokine receptors signal through terminate the cell cycle 559
intracellular tyrosine kinases (JAKs) and transcription
factors (STATs) 526 13.4 Sister chromatids in mitosis 560
The gp130 and γc subunits are common to many Cohesin holds sister chromatids together until
class I cytokine receptor assemblies 527 anaphase 561
 Detailed Contents xxiii

Interphase (G2) to metaphase is characterized by Chapter 14 Motility 597

chromosome condensation and then proteolytic
cleavage of cohesin 563 14.1 Introduction 597
13.5 Kinetochores in anaphase 564 14.2 Actin filaments and associated
Kinetochores are assembled on centromeric DNA proteins 598
associated with a novel histone, CENP-A 564 F-actin is a polar two-stranded filament 599
Kinetochores contain many proteins that form stable Actin filaments are dynamic: they grow and shorten,
complexes 565 coupled to ATP hydrolysis 599
Ndc80 from the KMN network is a crucial constituent The structure of G-actin was determined by X-ray
of the microtubule-binding interface 568 crystallography of co-crystals 600
Vertical and horizontal arrangements for the F-actin structure has been determined both by X-ray
microtubule-binding interface have been proposed 569 fiber diffraction and by reconstruction of
Motor proteins speed up the processes of cryo-electron micrographs 601
kinetochore/microtubule attachment 570 In F-actin, the strong inter-subunit bonds are in the
Polymerization and depolymerization of spindle axial direction 602
microtubules generates movement 570 In vivo, the structure and stability of filaments are
The spindle assembly checkpoint monitors the state of controlled by actin-binding proteins and can be
kinetochore/microtubule attachment before the affected by drugs 602
metaphase-to-anaphase transition is permitted 572 14.3 The myosin motor proteins 604
Conversion of O-Mad2 to C-Mad2 involves Myosin heavy chains have three domains—head,
templated refolding 574 neck, and tail 605
Kinetochores are able to distinguish between correct The first crystal structure of a myosin motor domain
and erroneous microtubule attachments 575 depicted subfragment 1 from chicken myosin II
Kinetochores are dynamic assemblies 576 without bound nucleotide 605
13.6 Centrioles and centrosomes 576 A four-state cycle describes ATP hydrolysis, actin
binding, and force generation 606
Centrosomes duplicate once per cell cycle 576
Myosin motors undergo nucleotide-dependent
Centriole duplication is controlled by a conserved conformational changes 607
set of proteins 579
The binding of myosin to actin and its binding of
Duplicated centriole pairs remain associated until nucleotides are mutually antagonistic 609
G2/M 580 In vitro motility assays support the swinging lever
Centrioles are sites of assembly of PCM 581 arm model of contraction 610
Microtubules are nucleated by the γ-tubulin ring The polarity of a myosin motor can be reversed by
complex 581 re-orienting the converter domain through genetic
The yeast SPB suggests a structure for the engineering 611
centrosome and microtubule initiation 582 The forces generated by individual myosin motors
The organization of the SPB proteins is focused and their step-lengths have been measured by
on Spc42 582 optical trapping 611
Centrosomes have other roles beyond cell division 584 14.4 Force generation in muscle 612
13.7 Apoptosis (programmed cell death) 585 Thick and thin filaments are packed in hexagonal
arrays 614
Apoptosis proceeds by intrinsic and extrinsic pathways 585
Muscles contract and force is generated by a sliding
Caspases mediate an intracellular proteolytic cascade 586 filament mechanism 614
Caspases bring about a multitude of changes in the Force generation is accompanied by structural
cell that lead ultimately to its death 587 changes in the myofibrils 616
Assembly of the apoptosome is critical to the Muscle achieves ~50% efficiency as a force-
intrinsic pathway 588 generating machine 617
Proteins of the Bcl-2 family are required for Muscles are switched on by excitation–contraction
cytochrome c release from mitochondria 589 coupling 617
The extrinsic pathway of apoptosis is mediated by Contraction of striated muscle is regulated by changes
membrane-associated DISCs 591 in the interactions of troponin and tropomyosin with
There is cross-talk between the pathways of actin filaments 619
apoptosis and other cellular pathways 592
14.5 Myosin filaments 619
The apoptosome of C. elegans presents a simple
mode of caspase activation 593 Myosin filaments have helical arrangements of
molecules 619
13.8 Summary 594 In relaxed thick filaments, there is regulatory cross-
References 595 talk between the two heads of a myosin molecule 620
xxiv Detailed Contents

Thick filaments contain accessory proteins with 14.10 Motility powered by

structural and regulatory roles 621 supramolecular springs 647
In smooth muscle, myosin filaments have a Contractile bacteriophage tails are dynamic
side-polar structure and their state of assembly gene-delivery systems 647
is regulated by phosphorylation 622 Fertilization by Limulus sperm involves uncoiling a
14.6 Microtubules and associated long bundle of cross-linked actin filaments 648
proteins 623 Tensed macromolecular springs drive other reactions
There is a discontinuity or ‘seam’ in the helical from insect hopping to membrane fusion 649
lattices of most microtubules 623 14.11 Chemotaxis I: the bacterial flagellum 649
Assembly, disassembly, and stability are influenced The rotary motor that drives flagellar motility is
by GTP hydrolysis 625 reversible and powered by ion gradients 650
Intracellular assembly of microtubules initiates in The flagellum has a modular architecture built from
specific nucleation complexes 626 24 different proteins 651
Uncapped microtubules grow and shorten at both The flagellar filament is a hollow tube with a
ends, asymmetrically 627 backbone of packed α helices 653
Dynamic instability allows microtubules to search for The hook functions as a universal joint and the rod
targets 627 serves as the drive shaft of the flagellar motor 654
The state of tubulin assembly can be altered by drug Six basal-body proteins contribute to torque generation 656
binding 628
The flagellar motor is a powerful and efficient
Destabilizing proteins cause microtubules to stepping motor 656
disassemble 629
Microtubule-associated proteins stabilize the polymers 631 14.12 Chemotaxis II: signaling by
chemoreceptor arrays 657
MAP activity is controlled by phosphorylation 632
Chemoreceptors modulate the activity of a kinase,
14.7 Motorized transport along CheA 658
microtubules 632 Chemoreceptors have globular periplasmic domains
Kinesin families have distinctive domain and coiled-coil cytoplasmic domains 658
arrangements 632 Chemotactic signals are conveyed by allosteric
Interaction of kinesin with microtubules is controlled switches 659
by the ATPase cycle of the motor domain 634 CheA is the engine that powers the chemical signaling
Most kinesin motors cannot operate singly but must events of the chemotaxis pathway 659
collaborate in groups 634 The receptor cluster is a plate-like assembly of
Kinesin movement is best explained by a Brownian ‘trimers-of-dimers’ 660
ratchet mechanism 635
14.13 Summary 661
The binding of motors can be visualized by cryo-EM
of decorated microtubules 635 References 661
Dynein is a ring-shaped molecule with six AAA+
ATPase domains 636
Chapter 15 Bioenergetics 663
The ATPase cycle of dynein powers minus
end-directed transport 637 15.1 Introduction 663
Cytoplasmic dynein has important roles at both 15.2 Biological oxidation and the
ends of microtubules 638 respiratory chain 665
14.8 Motile organelles built from The free energy released in the oxidation of NADH
microtubules 639 and FADH2 is stored as an electrochemical proton
The axonemal bundle of microtubules has a 9:2 gradient 666
symmetry mismatch 640 Complex I (NADH–ubiquinone oxidoreductase) is
Dynein heads are stacked in the outer and inner arms 640 the entry point for oxidation of NADH 670
Beating results from dynein-driven sliding of Structure of intact Complex I and a possible proton
adjacent microtubules 641 pumping mechanism 672
The mitochondrial Complex I resembles the bacterial
14.9 Polymerization/depolymerization Complex I but has many more subunits 674
machines 642 Complex II (succinate–ubiquinone oxidoreductase)
Brownian ratchets generate force by channeling is a succinate dehydrogenase and not a proton pump 675
random motions in particular directions 642 Complex III (cytochrome bc1 complex) oxidizes the
Tubulin, actin, and other proteins function as QH2 produced by Complexes I and II 676
polymerization engines 644 Proton pumping by Complex III depends on a
Cell migration is powered by polymerization engines 645 Q cycle 677
 Detailed Contents xxv

The ISPs in the Complex III dimer undergo large The structure of the F1 component reveals the rotary
structural movements 678 mechanism of ATP synthase 708
Complex IV (cytochrome c oxidase) reduces Single-molecule experiments demonstrate rotation
molecular oxygen to water 679 of the γ subunit in F1 710
Reduction of molecular oxygen is a four-electron The rotation of the γ subunit in F1 can be broken
reaction 680 down into steps 712
Protons enter and leave the membrane-embedded The c-subunits form a ring in Fo that rotates against a
CcO through gated pathways 680 stator complex 712
The oxidation of QH2 in some prokaryotes is Proton transport across the membrane by the
catalyzed by specialized quinol oxidases 682 c-ring makes it rotate 714
Respiratory chain complexes are assembled by The transport of protons can be correlated with the
modular pathways 682 efficiency of ATP synthesis 715
Respiratory chain complexes may come together in ATP synthase dimers help shape the mitochondrial
higher-order structures 684 cristae and form respiratory supercomplexes 716
Respiration can produce dangerous side-products, 15.5 Summary 717
which are eliminated by protective enzymes 685
References 718
The respiratory chain is very efficient in the capture
of energy as a proton-motive force 685
15.3 Photosynthetic reaction centers Chapter 16 Membrane Channels
and light-harvesting complexes 686 and Transporters 721
In plants the photosynthetic machinery is located
in organelles called chloroplasts 686 16.1 Introduction 721
Photosynthesis depends on the photochemical 16.2 Ion channels 721
capabilities of light-absorbing pigments 687 Ion channels have pores that are highly selective
Chlorophylls and carotenoids are crucial and gated 722
components of reaction centers and light-harvesting The KcsA channel allows a rapid and highly
(antenna) complexes 691 selective flux of potassium ions 722
Two types of LHC serve different purposes in purple Gating is mediated through extra domains attached
bacteria 692 to the pore unit 724
The LHCs of plants and cyanobacteria differ from Inward rectifier K+ (Kir) channels are regulated by
those of purple bacteria 693 cytoplasmic gating domains 724
Energy transfer reactions in LHCs are very fast Voltage gating requires a movement of charge
and very efficient 694 through the membrane 726
The structural blueprint of the RC of purple The glutamate receptor combines a large extracellular
photosynthetic bacteria is conserved throughout region with a conventional K+-selective pore 728
photosynthetic organisms 695
The trimeric acid-sensing ion channel (ASIC)
In purple bacteria, the cytochrome bc1 complex imposes Na+ selectivity by its pore architecture 728
generates a proton gradient that drives ATP synthesis 697
Chloride channels (CLCs) have conserved pore
In cyanobacteria and plant chloroplasts, two RCs regions and diverse extracellular and cytoplasmic
work in series with water as electron donor 697 regions 730
Photosystem II oxidizes water and reduces a quinone 698 The mechanosensitive ion channel MscS has a
The oxidation of water is a four-electron process heptameric structure 732
catalyzed by a specialized Mn cluster in photosystem II 699
16.3 Nicotinic acetylcholine receptor 733
A cytochrome b6 f complex links photosystem II to
photosystem I and generates a proton gradient 701 The ACh receptor is highly selective for cations 733
Photosystem I generates a reductant powerful The ACh receptor has evolved to be extremely
enough to reduce CO2 to carbohydrate 702 fast-acting and efficient 734
ATP can be generated by cyclic and non-cyclic The two ligand-binding sites have different affinities 735
electron transport 704 The gate is a hydrophobic girdle in the middle of
Photosynthesis is very efficient in the capture of the membrane 736
solar energy 705 Gating involves movements in the ligand-binding
and membrane-spanning domains 736
15.4 Electrochemical potential and the
biosynthesis of ATP 706 16.4 Aquaporins 737
The F1Fo-ATP synthase is constructed from two The structure of aquaporin 4 shows the basis for
rotary motors 706 water conductance 739
The F1 component functions by means of a chemical The wider selectivity filter of GlpF accounts for its
binding change mechanism 708 preference for glycerol 740
xxvi Detailed Contents

16.5 Transporters 741 17.3 The complement system 768

ABC transporters employ an alternating access The classical and lectin pathways use the multimeric
mechanism driven by ATP hydrolysis 741 complexes C1, mannose-binding lectin (MBL), and
The transmembrane domains exhibit a variety of the ficolins for pathogen recognition 769
helical arrangements 742 The first step in C1 activation involves C1q
The nucleotide-binding domains form dimers recognition of the target and auto-activation of C1r 771
with ATP 743 The alternative pathway and the classical and lectin
Major facilitator superfamily proteins tap into pathways converge on the C3 convertase 772
concentration gradients to transport their substrates 744 Cleavage of C3 causes major structural changes that
lead to activation 772
The transport mechanism of nucleobase cation
symport 1 exploits pseudosymmetry 747 Amplification of the response involves the C3
convertase, C3bBb 774
A multidrug resistance protein employs a rotary
version of the alternating access mechanism 748 Complement activation leads to four major
consequences: cell lysis, an inflammatory response,
16.6 The P-type ATPase pumps 750 phagocytosis, and B cell stimulation 777
The SERCA ATPase pumps Ca2+ out of the Host cells are protected by regulator proteins 778
cytoplasm 751 Several viruses and bacteria use host protection
The binding sites for Ca2+ and ATP in Ca2+-ATPase mechanisms to evade complement-mediated
are 50 Å apart 751 clearance 780
Conformational changes switch the enzyme between 17.4 T-cell-mediated immunity 780
different functional states 752
Major histocompatibility complexes (MHCs) are
The conformational change from E1P to E2P delivers glycoproteins that present foreign and self-antigens
Ca2+ to the lumen 753 at the surface of antigen-presenting cells (APCs) 783
A mechanism for Ca2+ transport is based on Peptides are loaded on to MHC-I and MHC-II via
alternating access 754 different pathways 785
The Na+/K+-ATPase regulates the cellular CD1 and other nonclassical MHC molecules may
concentrations of Na+ and K+ 755 have functions other than antigen presentation 786
The structure of Na+/K+-ATPase differs from The T cell receptor (TCR) is the primary antigen-
Ca2+-ATPase at the cation-binding sites 756 recognition molecule on the surface of T cells 786
16.7 Summary 757 The CD3 complex initiates and transmits signals in
the T cell interior 788
References 758
The TCR/CD3 complex: expression of the TCR at the
cell surface is accompanied by co-expression of CD3 789
Chapter 17 Complexes of the TCR co-receptor molecules are required for T cell
development and activation 790
Immune System 761 Antigen recognition by the TCR initiates intracellular
17.1 Introduction 761 signaling via the associated CD3 complex 791
Phosphorylation of CD3 leads to activation of T cells 791
17.2 Toll-like receptors and
inflammasomes 762 The immunological synapse mediates the cytolytic
machinery of cytotoxic T cells 793
Toll-like receptors invoke an inflammatory response
that also assists the adaptive immune response 762 17.5 Summary 795
Toll-like receptors recognize a variety of ligands References 796
within a common framework 763
The cytosolic TIR domains of Toll-like receptors
promote downstream signaling 765 Glossary 799
Pattern-recognition receptors promote the assembly
of inflammasomes 767 Index 817
 xxvii

Life Processes are Driven by

Macromolecular Assemblies and Machines

Life on Earth depends on two key features. One is the presence of water to enable the chemi-
cal reactions on which life depends. The second is a system for capturing information and
passing it on to succeeding generations. It is now clear that both features relate to the prop-
erties of macromolecular complexes. Water as a medium is paramount for them to assume
their native functional structures and to support sufficient mobility of the complexes and
their substrates in a fluid phase. A subset of these complexes is responsible for information
storage, expression, and replication.
For many years now, scientists have sought to understand life processes by studying indi-
vidually the many and varied macromolecules that make up a cell—the smallest viable unit
of life. This reductionist approach of isolating macromolecules for structural and functional
investigation has, in fact, been enormously successful. The atomic resolution structures of
DNA and proteins such as myoglobin, hemoglobin, and lysozyme in the 1950s and 1960s
were triumphs of the then-emerging discipline of structural molecular biology and pro-
vided a solid foundation for understanding their mechanisms of action. Nowadays, the
myriad solved structures in the Protein Data Bank (PDB, http://www.rcsb.org) as well as the
smaller but growing Electron Microscopy Data Bank (EMDB, http://www.ebi.ac.uk/pdbe/
emdb/) attest to the success of this approach and afford invaluable resources for molecular
biologists, biochemists, and life scientists generally.
In spite of these achievements, awareness has grown in recent years that only rarely can a
complex biological function be attributed to an individual macromolecule. Rather, most cel-
lular functions are performed by assemblies of several different species of macromolecules
(proteins, RNA, and DNA, as appropriate) acting in concert, and the ability of an assembly
to function depends critically on its structural integrity. In general, the structure and func-
tion of an assembly cannot be inferred from the properties of its components alone: the
scope of an assembly extends far beyond the sum of its parts. It is evident that close coor-
dination of components can greatly improve the speed and efficiency of its operation and
make possible reactions of staggering complexity (Figure 1). Many assemblies are endowed
with explicitly machine-like properties and they are the central theme of this book. Other
assemblies have primarily structural roles as ‘smart’ biomaterials but exhibit a similar diver-
sity in their composition and sophistication in how they are regulated.
In a seminal review published in 1998, Bruce Alberts wrote: “we now know that nearly every
major process in a cell is carried out by assemblies of 10 or more protein molecules. And, as
it carries out its biological functions, each of these protein assemblies interacts with several
other large complexes of proteins. Indeed, the entire cell can be viewed as a factory that
contains an elaborate network of interlocking assembly lines, each of which is composed of
a set of large protein machines.”
Assemblies are held together by weak (noncovalent) interactions and they vary widely in
stability and longevity. The methods used to break open cells to allow the isolation of their
contents may break down labile assemblies, and removal from the crowded environment
of a cell interior may have a similar effect. Complexes robust enough to withstand the rig-
ors of purification can be studied structurally with the traditional methods—primarily
X-ray crystallography and cryo-electron microscopy. However, labile complexes can be of
no less importance, and fleeting interactions can be crucially important to the cell. Frag-
ile complexes may be amenable to a ‘divide and conquer’ approach in which individual Figure 1 Charlie Chaplin and friend ponder
protein subunits or subcomplexes are solved at high resolution by X-ray crystallography the workings of a machine. (From Modern
or nuclear magnetic resonance (NMR) spectroscopy or computational methods, and then MBAM
Times. With MB1.1.01/p1
permission from Roy Export S.A.S.)
xxviii

fitted together to match the structure of the complex as determined by electron microscopy
(EM). Until recently, EM structures have been at lower resolution but technical advances
are now yielding—in favorable cases—structures of complexes and subcomplexes at resolu-
tions comparable to those achieved in X-ray crystallography. On a broader front, labile and
transient complexes may be pursued by ‘hybrid’ approaches that also integrate information
from methods such as chemical cross-linking, mass spectrometry, hydrodynamic analysis,
small-angle X-ray scattering, predictive bioinformatics, fluorescence techniques, and elec-
tron paramagnetic resonance spectroscopy.
Almost all major cellular functions are carried out by assemblies and it appears that almost
the entire content of a cell (other than water and ions) consists of large structures that can
be sedimented by centrifugation. However, it is difficult to obtain reliable estimates for the
numbers of assemblies—stable, unstable, or transient—in any given cell, even one as small
as a bacterium. Interaction maps derived from large-scale proteomics experiments provide a
glimpse of the abundance of some complexes, but most remain ill defined because it is noto-
riously difficult to discriminate between direct physical interactions and indirect functional
links, between binary and multiple interactions, and between transient (‘kiss and run’) and
stable interactions. It is possible, even likely, that many metabolic or signaling pathways
are organized in ways that are difficult to observe, by nonrandom interactions mediated by
weak intermolecular forces. To elucidate them—and to fully relate findings made in vitro
with what happens inside a cell—will require the further development and integration of
methods such as electron tomography and ‘super-resolution’ light microscopy that allow
structural studies in situ; that is, in unperturbed cellular environments. Thus, structural
biology and cell biology need to converge to achieve a deeper understanding of the multi-
faceted organization of cells. That is the challenge for the emerging discipline of mesobiol-
ogy: to rationalize the biological activity of macromolecular assemblies and machines.

References
Alberts B (1998) The cell as a collection of protein machines: preparing the next generation of
molecular biologists. Cell 92:291–294.
Ball P (2003) Portrait of a molecule. Nature 421:421–422.
Campbell ID (2008) Structure of the living cell. Phil Trans Roy Soc B 363:2379–2391.
Hartwell LH, Hopfield JL, Leibler S & Murray AW (1999) From molecular to modular cell biology.
Nature 402:C47–C52 (6761 Suppl.).
Gierasch LM & Gershenson A (2009) Post-reductionist protein science, or putting Humpty Dumpty
back together again. Nat Chem Biol 5:774–777.
Sali A, Glaeser R, Earnest T & Baumeister W (2003) From words to literature in structural
proteomics. Nature 422:216–225.
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matter. In 1816, I had not acquiesced in the tariff, then supported by
South Carolina. To some parts of it, especially, I felt and expressed
great repugnance. I held the same opinions in 1821, at the meeting in
Faneuil Hall, to which the gentleman has alluded. I said then, and
say now, that, as an original question, the authority of Congress to
exercise the revenue power, with direct reference to the protection of
manufactures, is a questionable authority, far more questionable in
my judgment, than the power of internal improvements. I must
confess, sir, that, in one respect, some impression has been made on
my opinions lately. Mr. Madison’s publication has put the power in a
very strong light. He has placed it, I must acknowledge, upon
grounds of construction and argument which seem impregnable. But
even if the power were doubted, on the face of the constitution itself,
it had been assumed and asserted in the first revenue law ever
passed under the same constitution; and, on this ground, as a matter
settled by contemporaneous practice, I had refrained from
expressing the opinion that the tariff laws transcended constitutional
limits, as the gentleman supposes. What I did say at Faneuil Hall, as
far as I now remember, was, that this was originally matter of
doubtful construction. The gentleman himself, I suppose, thinks
there is no doubt about it, and that the laws are plainly against the
constitution. Mr. Madison’s letters, already referred to, contain, in
my judgment, by far the most able exposition extant of this part of
the constitution. He has satisfied me, so far as the practice of the
government had left it an open question.
With a great majority of the representatives of Massachusetts, I
voted against the tariff of 1824. My reasons were then given, and I
will not now repeat them. But notwithstanding our dissent, the great
states of New York, Pennsylvania, Ohio, and Kentucky went for the
bill, in almost unbroken column, and it passed. Congress and the
president sanctioned it, and it became the law of the land. What,
then, were we to do? Our only option was either to fall in with this
settled course of public policy, and to accommodate ourselves to it as
well as we could, or to embrace the South Carolina doctrine, and talk
of nullifying the statute by state interference.
The last alternative did not suit our principles, and, of course, we
adopted the former. In 1827, the subject came again before Congress,
on a proposition favorable to wool and woolens. We looked upon the
system of protection as being fixed and settled. The law of 1824
remained. It had gone into full operation, and in regard to some
objects intended by it, perhaps most of them had produced all its
expected effects. No man proposed to repeal it—no man attempted to
renew the general contest on its principle. But, owing to subsequent
and unforeseen occurrences, the benefit intended by it to wool and
woolen fabrics had not been realized. Events, not known here when
the law passed, had taken place, which defeated its object in that
particular respect. A measure was accordingly brought forward to
meet this precise deficiency, to remedy this particular defect. It was
limited to wool and woolens. Was ever any thing more reasonable? If
the policy of the tariff laws had become established in principle as
the permanent policy of the government, should they not be revised
and amended, and made equal, like other laws, as exigencies should
arise, or justice require? Because we had doubted about adopting the
system, were we to refuse to cure its manifest defects after it became
adopted, and when no one attempted its repeal? And this, sir, is the
inconsistency so much bruited. I had voted against the tariff of 1824
—but it passed; and in 1827 and 1828, I voted to amend it in a point
essential to the interest of my constituents. Where is the
inconsistency? Could I do otherwise?
Sir, does political consistency consist in always giving negative
votes? Does it require of a public man to refuse to concur in
amending laws because they passed against his consent? Having
voted against the tariff originally, does consistency demand that I
should do all in my power to maintain an unequal tariff, burdensome
to my own constituents, in many respects,—favorable in none? To
consistency of that sort I lay no claim; and there is another sort to
which I lay as little—and that is, a kind of consistency by which
persons feel themselves as much bound to oppose a proposition after
it has become the law of the land as before.
The bill of 1827, limited, as I have said, to the single object in
which the tariff of 1824 had manifestly failed in its effects, passed the
House of Representatives, but was lost here. We had then the act of
1828. I need not recur to the history of a measure so recent. Its
enemies spiced it with whatsoever they thought would render it
distasteful; its friends took it, drugged as it was. Vast amounts of
property, many millions, had been invested in manufactures, under
the inducements of the act of 1824. Events called loudly, I thought,
for further regulations to secure the degree of protection intended by
that act. I was disposed to vote for such regulations and desired
nothing more; but certainly was not to be bantered out of my
purpose by a threatened augmentation of duty on molasses, put into
the bill for the avowed purpose of making it obnoxious. The vote may
have been right or wrong, wise or unwise; but it is a little less than
absurd to allege against it an inconsistency with opposition to the
former law.
Sir, as to the general subject of the tariff, I have little now to say.
Another opportunity may be presented. I remarked, the other day,
that this policy did not begin with us in New England; and yet, sir,
New England is charged with vehemence as being favorable, or
charged with equal vehemence as being unfavorable, to the tariff
policy, just as best suits the time, place, and occasion for making
some charge against her. The credulity of the public has been put to
its extreme capacity of false impression relative to her conduct in this
particular. Through all the south, during the late contest, it was New
England policy, and a New England administration, that was
inflicting the country with a tariff policy beyond all endurance, while
on the other side of the Alleghany, even the act of 1828 itself—the
very sublimated essence of oppression, according to southern
opinions—was pronounced to be one of those blessings for which the
west was indebted to the “generous south.”
With large investments in manufacturing establishments, and
various interests connected with and dependent on them, it is not to
be expected that New England, any more than other portions of the
country, will now consent to any measures destructive or highly
dangerous. The duty of the government, at the present moment,
would seem to be to preserve, not to destroy; to maintain the
position which it has assumed; and for one, I shall feel it an
indispensable obligation to hold it steady, as far as in my power, to
that degree of protection which it has undertaken to bestow. No
more of the tariff.
Professing to be provoked by what he chose to consider a charge
made by me against South Carolina, the honorable member, Mr.
President, has taken up a new crusade against New England. Leaving
altogether the subject of the public lands, in which his success,
perhaps, had been neither distinguished nor satisfactory, and letting
go, also, of the topic of the tariff, he sallied forth in a general assault
on the opinions, politics, and parties of New England, as they have
been exhibited in the last thirty years. This is natural. The “narrow
policy” of the public lands had proved a legal settlement in South
Carolina, and was not to be removed. The “accursed policy” of the
tariff, also, had established the fact of its birth and parentage in the
same state. No wonder, therefore, the gentleman wished to carry the
war, as he expressed it, into the enemy’s country. Prudently willing
to quit these subjects, he was doubtless desirous of fastening others,
which could not be transferred south of Mason and Dixon’s line. The
politics of New England became his theme; and it was in this part of
his speech, I think, that he menaced me with such sore discomfiture.
Discomfiture! why, sir, when he attacks anything which I
maintain, and overthrows it; when he turns the right or left of any
position which I take up; when he drives me from any ground I
choose to occupy, he may then talk of discomfiture, but not till that
distant day. What has he done? Has he maintained his own charges?
Has he proved what he alleged? Has he sustained himself in his
attack on the government, and on the history of the north, in the
matter of the public lands? Has he disproved a fact, refuted a
proposition, weakened an argument maintained by me? Has he come
within beat of drum of any position of mine? O, no; but he has
“carried the war into the enemy’s country!” Carried the war into the
enemy’s country! Yes, sir, and what sort of a war has he made of it?
Why, sir, he has stretched a dragnet over the whole surface of
perished pamphlets, indiscreet sermons, frothy paragraphs, and
fuming popular addresses; over whatever the pulpit in its moments
of alarm, the press in its heats, and parties in their extravagances,
have severally thrown off, in times of general excitement and
violence. He has thus swept together a mass of such things, as, but
they are not now old, the public health would have required him
rather to leave in their state of dispersion.
For a good long hour or two, we had the unbroken pleasure of
listening to the honorable member, while he recited, with his usual
grace and spirit, and with evident high gusto, speeches, pamphlets,
addresses, and all that et ceteras of the political press, such as warm
heads produce in warm times, and such as it would be “discomfiture”
indeed for any one, whose taste did not delight in that sort of
reading, to be obliged to peruse. This is his war. This is to carry the
war into the enemy’s country. It is in an invasion of this sort that he
flatters himself with the expectation of gaining laurels fit to adorn a
senator’s brow.
Mr. President, I shall not, it will, I trust, not be expected that I
should, either now or at any time, separate this farrago into parts,
and answer and examine its components. I shall hardly bestow upon
it all a general remark or two. In the run of forty years, sir, under this
constitution, we have experienced sundry successive violent party
contests. Party arose, indeed, with the constitution itself, and in
some form or other has attended through the greater part of its
history.
Whether any other constitution than the old articles of
confederation was desirable, was itself, a question on which parties
divided; if a new constitution was framed, what powers should be
given to it was another question; and when it had been formed, what
was, in fact, the just extent of the powers actually conferred was a
third. Parties, as we know, existed under the first administration, as
distinctly marked as those which manifested themselves at any
subsequent period.
The contest immediately preceding the political change in 1801,
and that, again, which existed at the commencement of the late war,
are other instances of party excitement, of something more than
usual strength and intensity. In all these conflicts there was, no
doubt, much of violence on both and all sides. It would be
impossible, if one had a fancy for such employment, to adjust the
relative quantum of violence between these two contending parties.
There was enough in each, as must always be expected in popular
governments. With a great deal of proper and decorous discussion
there was mingled a great deal, also, of declamation, virulence,
crimination, and abuse.
In regard to any party, probably, at one of the leading epochs in
the history of parties, enough may be found to make out another
equally inflamed exhibition as that with which the honorable
member has edified us. For myself, sir, I shall not rake among the
rubbish of by-gone times to see what I can find or whether I cannot
find something by which I can fix a blot on the escutcheon of any
state, any party, or any part of the country. General Washington’s
administration was steadily and zealously maintained, as we all
know, by New England. It was violently opposed elsewhere. We
know in what quarter he had the most earnest, constant and
persevering support, in all his great and leading measures. We know
where his private and personal character was held in the highest
degree of attachment and veneration; and we know, too, where his
measures were opposed, his services slighted, and his character
vilified.
We know, or we might know, if we turn to the journals, who
expressed respect, gratitude, and regret, when he retired from the
chief magistracy; and who refused to express either respect,
gratitude or regret. I shall not open those journals. Publications more
abusive or scurrilous never saw the light than were sent forth against
Washington, and all his leading measures, from presses south of New
England; but I shall not look them up. I employ no scavengers—no
one is in attendance on me, tendering such means of retaliation; and
if there were, with an ass’s load of them, with a bulk as huge as that
which the gentleman himself has produced, I would not touch one of
them. I see enough of the violence of our own times to be no way
anxious to rescue from forgetfulness the extravagances of times past.
Besides, what is all this to the present purpose? It has nothing to do
with the public lands, in regard to which the attack was begun; and it
has nothing to do with those sentiments and opinions, which I have
thought tend to disunion, and all of which the honorable member
seems to have adopted himself, and undertaken to defend. New
England has, at times—so argues the gentleman,—held opinions as
dangerous as those which he now holds. Be it so. But why, therefore,
does he abuse New England? If he finds himself countenanced by
acts of hers, how is it that, while he relies on these acts, he covers, or
seeks to cover, their authors with reproach?
But, sir, if, in the course of forty years, there have been undue
effervescences of party in New England, has the same thing
happened no where else? Party animosity and party outrage, not in
New England, but elsewhere, denounced President Washington, not
only as a federalist, but as a tory, a British agent, a man who, in his
high office, sanctioned corruption. But does the honorable member
suppose that, if I had a tender here, who should put such an effusion
of wickedness and folly in my hand, that I would stand up and read it
against the south? Parties ran into great heats, again, in 1799. What
was said, sir, or rather what was not said, in those years, against
John Adams, one of the signers of the Declaration of Independence,
and its admitted ablest defender on the floor of Congress? If the
gentleman wants to increase his stores of party abuse and frothy
violence, if he has a determined proclivity to such pursuits, there are
treasures of that sort south of the Potomac, much to his taste, yet
untouched. I shall not touch them.
The parties which divided the country, at the commencement of
the late war, were violent. But, then, there was violence on both
sides, and violence in every state. Minorities and majorities were
equally violent. There was no more violence against the war in New
England than in other states; nor any more appearance of violence,
except that, owing to a dense population, greater facility for
assembling, and more presses, there may have been more, in
quantity, spoken and printed there than in some other places. In the
article of sermons, too, New England is somewhat more abundant
than South Carolina: and for that reason, the chance of finding here
and there an exceptionable one may be greater. I hope, too, there are
more good ones. Opposition may have been more formidable in New
England, as it embraced a larger portion of the whole population: but
it was no more unrestrained in its principle, or violent in manner.
The minorities dealt quite as harshly with their own state
governments as the majorities dealt with the administration here.
There were presses on both sides, popular meetings on both sides,
ay, and pulpits on both sides, also. The gentleman’s purveyors have
only catered for him among the productions of one side. I certainly
shall not supply the deficiency by furnishing samples of the other. I
leave to him, and to them, the whole concern.
It is enough for me to say, that if, in any part of this, their grateful
occupation—if in all their researches—they find anything in the
history of Massachusetts, or New England, or in the proceedings of
any legislative or other public body, disloyal to the Union, speaking
slightly of its value, proposing to break it up, or recommending non-
intercourse with neighboring states, on account of difference of
political opinion, then, sir, I give them all up to the honorable
gentleman’s unrestrained rebuke; expecting, however, that he will
extend his buffetings, in like manner, to all similar proceedings,
wherever else found.
The gentleman, sir, has spoken at large of former parties, now no
longer in being, by their received appellations, and has undertaken to
instruct us, not only in the knowledge of their principles, but of their
respective pedigrees also. He has ascended to their origin and run
out their genealogies. With most exemplary modesty, he speaks of
the party to which he professes to have belonged himself, as the true,
pure, the only honest, patriotic party, derived by regular descent,
from father to son, from the time of the virtuous Romans! Spreading
before us the family tree of political parties, he takes especial care to
show himself snugly perched on a popular bough! He is wakeful to
the expediency of adopting such rules of descent, for political parties,
as shall bring him in, in exclusion of others, as an heir to the
inheritance of all public virtue, and all true political principles. His
doxy is always orthodoxy. Heterodoxy is confined to his opponents.
He spoke, sir, of the federalists, and I thought I saw some eyes begin
to open and stare a little, when he ventured on that ground. I
expected he would draw his sketches rather lightly, when he looked
on the circle round him, and especially if he should cast his thoughts
to the high places out of the Senate. Nevertheless, he went back to
Rome, ad annum urbs condita, and found the fathers of the
federalists in the primeval aristocrats of that renowned empire! He
traced the flow of federal blood down through successive ages and
centuries, till he got into the veins of the American tories, (of whom,
by the way, there were twenty in the Carolinas for one in
Massachusetts.) From the tories, he followed it to the federalists; and
as the federal party was broken up, and there was no possibility of
transmitting it farther on this side of the Atlantic, he seems to have
discovered that it has gone off, collaterally, though against all the
canons of descent, into the ultras of France, and finally became
extinguished, like exploded gas, among the adherents of Don Miguel.
This, sir, is an abstract of the gentleman’s history of federalism. I
am not about to controvert it. It is not, at present, worth the pains of
refutation, because, sir, if at this day one feels the sin of federalism
lying heavily on his conscience, he can easily obtain remission. He
may even have an indulgence, if he is desirous of repeating the
transgression. It is an affair of no difficulty to get into this same right
line of patriotic descent. A man, nowadays, is at liberty to choose his
political parentage. He may elect his own father. Federalist or not, he
may, if he choose, claim to belong to the favored stock, and his claim
will be allowed. He may carry back his pretensions just as far as the
honorable gentleman himself; nay, he may make himself out the
honorable gentleman’s cousin, and prove satisfactorily that he is
descended from the same political great-grandfather. All this is
allowable. We all know a process, sir, by which the whole Essex
Junto could, in one hour be all washed white from their ancient
federalism, and come out every one of them, an original democrat,
dyed in the wool! Some of them have actually undergone the
operation, and they say it is quite easy. The only inconvenience it
occasions, as they tell us, is a slight tendency of the blood to the face,
a soft suffusion, which, however, is very transient, since nothing is
said calculated to deepen the red on the cheek, but a prudent silence
observed in regard to all the past. Indeed, sir, some smiles of
approbation have been bestowed, and some crumbs of comfort have
fallen, not a thousand miles from the door of the Hartford
Convention itself. And if the author of the ordinance of 1787
possessed the other requisite qualifications, there is no knowing,
notwithstanding his federalism, to what heights of favor he might not
yet attain.
Mr. President, in carrying his warfare, such as it was, into New
England, the honorable gentleman all along professes to be acting on
the defensive. He desires to consider me as having assailed South
Carolina and insists that he comes forth only as her champion, and
in her defence. Sir, I do not admit that I made any attack whatever
on South Carolina. Nothing like it. The honorable member, in his
first speech, expressed opinions, in regard to revenue, and some
other topics, which I heard both with pain and surprise. I told the
gentleman that I was aware that such sentiments were entertained
OUT of the government, but had not expected to find them advanced
in it; that I knew there were persons in the south who speak of our
Union with indifference, or doubt, taking pains to magnify its evils,
and to say nothing of its benefits; that the honorable member
himself, I was sure, could never be one of these; and I regretted the
expression of such opinions as he had avowed, because I thought
their obvious tendency was to encourage feelings of disrespect to the
Union, and to weaken its connection. This, sir, is the sum and
substance of all I said on the subject. And this constitutes the attack
which called on the chivalry of the gentleman, in his opinion, to
harry us with such a forage among the party pamphlets and party
proceedings of Massachusetts. If he means that I spoke with
dissatisfaction or disrespect of the ebullitions of individuals in South
Carolina, it is true. But, if he means that I had assailed the character
of the state, her honor, or patriotism, that I had reflected on her
history or her conduct, he had not the slightest ground for any such
assumption. I did not even refer, I think, in my observations, to any
collection of individuals. I said nothing of the recent conventions. I
spoke in the most guarded and careful manner, and only expressed
my regret for the publication of opinions which I presumed the
honorable member disapproved as much as myself. In this, it seems,
I was mistaken.
I do not remember that the gentleman has disclaimed any
sentiment, or any opinion, of a supposed anti-Union tendency, which
on all or any of the recent occasions has been expressed. The whole
drift of his speech has been rather to prove, that, in divers times and
manners, sentiments equally liable to objection have been
promulgated in New England. And one would suppose that his
object, in this reference to Massachusetts, was to find a precedent to
justify proceedings in the south, were it not for the reproach and
contumely with which he labors, all along, to load his precedents.
By way of defending South Carolina from what he chooses to think
an attack on her, he first quotes the example of Massachusetts, and
then denounces that example, in good set terms. This twofold
purpose, not very consistent with itself, one would think, was
exhibited more than once in the course of his speech. He referred, for
instance, to the Hartford Convention. Did he do this for authority, or
for a topic of reproach? Apparently for both; for he told us that he
should find no fault with the mere fact of holding such a convention,
and considering and discussing such questions as he supposes were
then and there discussed; but what rendered it obnoxious was the
time it was holden, and the circumstances of the country then
existing. We were in a war, he said, and the country needed all our
aid; the hand of government required to be strengthened, not
weakened; and patriotism should have postponed such proceedings
to another day. The thing itself, then, is a precedent; the time and
manner of it, only, subject of censure.
Now, sir, I go much farther, on this point, than the honorable
member. Supposing, as the gentleman seems to, that the Hartford
Convention assembled for any such purpose as breaking up the
Union, because they thought unconstitutional laws had been passed,
or to concert on that subject, or to calculate the value of the Union;
supposing this to be their purpose, or any part of it, then I say the
meeting itself was disloyal, and obnoxious to censure, whether held
in time of peace, or time of war, or under whatever circumstances.
The material matter is the object. Is dissolution the object? If it be,
external circumstances may make it a more or less aggravated case,
but cannot affect the principle. I do not hold, therefore, that the
Hartford Convention was pardonable, even to the extent of the
gentleman’s admission, if its objects were really such as have been
imputed to it. Sir, there never was a time, under any degree of
excitement, in which the Hartford Convention, or any other
convention, could maintain itself one moment in New England, if
assembled for any such purpose as the gentleman says would have
been an allowable purpose. To hold conventions to decide questions
of constitutional law! to try the validity of statutes, by votes in a
convention! Sir, the Hartford Convention, I presume, would not
desire that the honorable gentleman should be their defender or
advocate, if he puts their case upon such untenable and extravagant
grounds.
Then, sir, the gentleman has no fault to find with these recently-
promulgated South Carolina opinions. And, certainly, he need have
none; for his own sentiments, as now advanced, and advanced on
reflection, as far as I have been able to comprehend them, go the full
length of all these opinions. I propose, sir, to say something on these,
and to consider how far they are just and constitutional. Before doing
that, however, let me observe, that the eulogium pronounced on the
character of the state of South Carolina, by the honorable gentleman,
for her revolutionary and other merits, meets my hearty
concurrence. I shall not acknowledge that the honorable member
goes before me in regard for whatever of distinguished talent or
distinguished character South Carolina has produced. I claim part of
the honor, I partake in the pride, of her great names. I claim them for
countrymen, one and all. The Laurenses, the Rutledges, the
Pinckneys, the Sumpters, the Marions—Americans all—whose fame
is no more to be hemmed in by state lines than their talents and their
patriotism were capable of being circumscribed within the same
narrow limits. In their day and generation, they served and honored
the country, and the whole country; and their renown is of the
treasures of the whole country. Him whose honored name the
gentleman himself bears—does he suppose me less capable of
gratitude for his patriotism, or sympathy for his sufferings, than if
his eyes had first opened upon the light in Massachusetts instead of
South Carolina? Sir, does he suppose it is in his power to exhibit a
Carolina name so bright as to produce envy in my bosom? No, sir,
increased gratification and delight, rather.
Sir, I thank God that if I am gifted with little of the spirit which is
said to be able to raise mortals to the skies, I have yet none, as I trust,
of that other spirit, which would drag angels down. When I shall be
found, sir, in my place here in the Senate, or elsewhere, to sneer at
public merit, because it happened to spring up beyond the little
limits of my own state, or neighborhood; when I refuse, for any such
cause, or for any cause, the homage due to American talent, to
elevated patriotism, to sincere devotion to liberty and the country; or
if I see an uncommon endowment of Heaven, if I see extraordinary
capacity and virtue in any son of the south, and if, moved by local
prejudice, or gangrened by state jealousy, I get up here to abate the
tithe of a hair from his just character and just fame,—may my tongue
cleave to the roof of my mouth! Sir, let me recur to pleasing
recollections; let me indulge in refreshing remembrance of the past;
let me remind you that in early times no states cherished greater
harmony, both of principle and feeling, than Massachusetts and
South Carolina. Would to God that harmony might again return.
Shoulder to shoulder they went through the revolution; hand in hand
they stood round the administration of Washington, and felt his own
great arm lean on them for support. Unkind feeling, if it exist,
alienation, and distrust are the growth, unnatural to such soils, of
false principles since sown. They are weeds, the seeds of which that
same great arm never scattered.
Mr. President, I shall enter on no encomium upon Massachusetts
—she needs none. There she is—behold her, and judge for yourselves.
There is her history—the world knows it by heart. The past, at least,
is secure. There is Boston, and Concord, and Lexington, and Bunker
Hill; and there they will remain forever. The bones of her sons, fallen
in the great struggle for independence, now lie mingled with the soil
of every state from New England to Georgia; and there they will lie
forever. And, sir, where American liberty raised its first voice, and
where its youth was nurtured and sustained, there it still lives, in the
strength of its manhood, and full of its original spirit. If discord and
disunion shall wound it; if folly and madness, if uneasiness under
salutary and necessary restraint, shall succeed to separate it from
that Union by which alone its existence is made sure,—it will stand,
in the end, by the side of that cradle in which its infancy was rocked;
it will stretch forth its arm, with whatever vigor it may still retain,
over the friends who gather around it; and it will fall at last, if fall it
must, amidst the proudest monuments of its glory, and on the very
spot of its origin.
There yet remains to be performed, Mr. President, by far the most
grave and important duty; which I feel to be devolved on me by this
occasion. It is to state, and to defend, what I conceive to be the true
principles of the constitution under which we are here assembled. I
might well have desired that so weighty a task should have fallen into
other and abler hands. I could have wished that it should have been
executed by those whose character and experience give weight and
influence to their opinions, such as cannot possibly belong to mine.
But, sir, I have met the occasion, not sought it; and I shall proceed to
state my own sentiments, without challenging for them any
particular regard, with studied plainness and as much precision as
possible.
I understand the honorable gentleman from South Carolina to
maintain that it is a right of the state legislatures to interfere,
whenever in their judgment, this government transcends its
constitutional limits, and to arrest the operation of its laws.
I understand him to maintain this right as a right existing under
the constitution, not as a right to overthrow it, on the ground of
extreme necessity, such as would justify violent revolution.
I understand him to maintain an authority, on the part of the
states, thus to interfere for the purpose of correcting the exercise of
power by the general government, of checking it, and of compelling it
to conform to their opinion of the extent of its power.
I understand him to maintain that the ultimate power of judging of
the constitutional extent of its own authority is not lodged
exclusively in the general government or any branch of it; but that,
on the contrary, the states may lawfully decide for themselves, and
each state for itself, whether, in a given case, the act of the general
government transcends its power.
I understand him to insist that, if the exigency of the case, in the
opinion of any state government, require it, such state government
may, by its own sovereign authority, annul an act of the general
government which it deems plainly and palpably unconstitutional.
This is the sum of what I understand from him to be the South
Carolina doctrine. I propose to consider it, and to compare it with the
constitution. Allow me to say, as a preliminary remark, that I call this
the South Carolina doctrine, only because the gentleman himself has
so denominated it. I do not feel at liberty to say that South Carolina,
as a state, has ever advanced these sentiments. I hope she has not,
and never may. That a great majority of her people are opposed to
the tariff laws is doubtless true. That a majority, somewhat less than
that just mentioned, conscientiously believe these laws
unconstitutional, may probably be also true. But that any majority
holds to the right of direct state interference, at state discretion, the
right of nullifying acts of Congress by acts of state legislation, is more
than I know, and what I shall be slow to believe.
That there are individuals, besides the honorable gentleman, who
do maintain these opinions, is quite certain. I recollect the recent
expression of a sentiment which circumstances attending its
utterance and publication justify us in supposing was not
unpremeditated—“The sovereignty of the state; never to be
controlled, construed, or decided on, but by her own feelings of
honorable justice.”
[Mr. Hayne here rose, and said, that for the purpose of being
clearly understood, he would state that his proposition was in the
words of the Virginia resolution, as follows:—
“That this Assembly doth explicitly and peremptorily declare, that it views the
powers of the federal government, as resulting from the compact, to which the
states are parties, as limited by the plain sense and intention of the instrument
constituting that compact, as no further valid than they are authorized by the
grants enumerated in that compact; and that, in case of a deliberate, palpable, and
dangerous exercise of other powers not granted by the same compact, the states
who are parties thereto have the right and are in duty bound, to interpose for
arresting the progress of the evil, and for maintaining, within their respective
limits, the authorities, rights, and liberties pertaining to them.”]
Mr. Webster resumed:—
I am quite aware, Mr. President, of the existence of the resolution
which the gentleman read, and has now repeated, and that he relies
on it as his authority. I know the source, too, from which it is
understood to have proceeded. I need not say, that I have much
respect for the constitutional opinions of Mr. Madison; they would
weigh greatly with me, always. But, before the authority of his
opinion be vouched for the gentleman’s proposition, it will be proper
to consider what is the fair interpretation of that resolution, to which
Mr. Madison is understood to have given his sanction. As the
gentleman construes it, it is an authority for him. Possibly he may
not have adopted the right construction. That resolution declares,
that in the case of the dangerous exercise of powers not granted by
the general government, the states may interpose to arrest the
progress of the evil. But how interpose? and what does this
declaration purport? Does it mean no more than that there may be
extreme cases in which the people, in any mode of assembling, may
resist usurpation, and relieve themselves from a tyrannical
government? No one will deny this. Such resistance is not only
acknowledged to be just in America, but in England also. Blackstone
admits as much, in the theory and practice, too, of the English
constitution. We, sir, who oppose the Carolina doctrine, do not deny
that the people may, if they choose, throw off any government, when
it becomes oppressive and intolerable, and erect a better in its stead.
We all know that civil institutions are established for the public
benefit, and that, when they cease to answer the ends of their
existence they may be changed.
But I do not understand the doctrine now contended for to be that
which, for the sake of distinctness, we may call the right of
revolution. I understand the gentleman to maintain, that without
revolution, without civil commotion, without rebellion, a remedy for
supposed abuse and transgression of the powers of the general
government lies in a direct appeal to the interference of the state
governments. [Mr. Hayne here rose: He did not contend, he said, for
the mere right of revolution, but for the right of constitutional
resistance. What he maintained was, that, in case of a plain, palpable
violation of the constitution by the general government, a state may
interpose; and that this interposition is constitutional.]
Mr. Webster resumed:
So, sir, I understood the gentleman, and am happy to find that I
did not misunderstand him. What he contends for is, that it is
constitutional to interrupt the administration of the constitution
itself, in the hands of those who are chosen and sworn to administer
it, by the direct interference, in form of law, of the states, in virtue of
their sovereign capacity. The inherent right in the people to reform
their government I do not deny; and that they have another right,
and that is, to resist unconstitutional laws without overturning the
government. It is no doctrine of mine, that unconstitutional laws
bind the people. The great question is, Whose prerogative is it to
decide on the constitutionality or unconstitutionality of the laws?
On that the main debate hinges. The proposition that, in the case of a
supposed violation of the constitution by Congress, the states have a
constitutional right to interfere, and annul the law of Congress, is the
proposition of the gentleman; I do not admit it. If the gentleman had
intended no more than to assert the right of revolution for justifiable
cause, he would have said only what all agree to.—But I cannot
conceive that there can be a middle course between submission to
the laws, when regularly pronounced constitutional, on the one
hand, and open resistance, which is revolution or rebellion, on the
other. I say the right of a state to annul a law of Congress cannot be
maintained but on the ground of the unalienable right of man to
resist oppression; that is to say, upon the ground of revolution. I
admit that there is no ultimate violent remedy, above the
constitution, and defiance of the constitution, which may be resorted
to, when a revolution is to be justified. But I do not admit that under
the constitution, and in conformity with it, there is any mode in
which a state government, as a member of the Union can interfere
and stop the progress of the general government, by force of her own
laws, under any circumstances whatever.
This leads us to inquire into the origin of this government, and the
source of its power. Whose agent is it? Is it the creature of the state
legislatures, or the creature of the people? If the government of the
United States be the agent of the state governments, then they may
control it, provided they can agree in the manner of controlling it; if
it is the agent of the people, then the people alone can control it,
restrain it, modify or reform it. It is observable enough, that the
doctrine for which the honorable gentleman contends leads him to
the necessity of maintaining, not only that this general government is
the creature of the states, but that it is the creature of each of the
states severally; so that each may assert the power, for itself, of
determining whether it acts within the limits of its authority. It is the
servant of four and twenty masters, of different wills and different
purposes; and yet bound to obey all. This absurdity (for it seems no
less) arises from a misconception as to the origin of this government,
and its true character. It is, sir, the people’s constitution, the people’s
government; made for the people; made by the people; and
answerable to the people. The people of the United States have
declared that this constitution shall be the supreme law. We must
either admit the proposition, or dispute their authority. The states
are unquestionably sovereign, so far as their sovereignty is not
affected by this supreme law. The state legislatures, as political
bodies, however sovereign, are yet not sovereign over the people. So
far as the people have given power to the general government, so far
the grant is unquestionably good, and the government holds of the
people, and not of the state governments. We are all agents of the
same supreme power, the people. The general government and the
state governments derive their authority from the same source.
Neither can, in relation to the other, be called primary; though one is
definite and restricted, and the other general and residuary.
The national government possesses those powers which it can be
shown the people have conferred on it, and no more. All the rest
belongs to the state governments, or to the people themselves. So far
as the people have restrained state sovereignty by the expression of
their will, in the constitution of the United States, so far, it must be
admitted, state sovereignty is effectually controlled. I do not contend
that it is, or ought to be, controlled further. The sentiment to which I
have referred propounds that state sovereignty is only to be
controlled by its own “feelings of justice;” that is to say, it is not to be
controlled at all; for one who is to follow his feelings, is under no
legal control. Now, however men may think this ought to be, the fact
is, that the people of the United States have chosen to impose control
on state sovereignties. The constitution has ordered the matter
differently from what this opinion announces. To make war, for
instance, is an exercise of sovereignty; but the constitution declares
that no state shall make war. To coin money is another exercise of
sovereign power; but no state is at liberty to coin money. Again: the
constitution says, that no sovereign state shall be so sovereign as to
make a treaty. These prohibitions, it must be confessed, are a control
on the state sovereignty of South Carolina, as well as of the other
states, which does not arise “from feelings of honorable justice.”
Such an opinion, therefore, is in defiance of the plainest provisions of
the constitution.
There are other proceedings of public bodies which have already
been alluded to, and to which I refer again for the purpose of
ascertaining more fully what is the length and breadth of that
doctrine, denominated the Carolina doctrine, which the honorable
member has now stood up on this floor to maintain.
In one of them I find it resolved that “the tariff of 1828, and every
other tariff designed to promote one branch of industry at the
expense of others, is contrary to the meaning and intention of the
federal compact; and as such a dangerous, palpable, and deliberate
usurpation of power, by a determined majority, wielding the general
government beyond the limits of its delegated powers, as calls upon
the states which compose the suffering minority, in their sovereign
capacity, to exercise the powers which, as sovereigns, necessarily
devolve upon them, when their compact is violated.”
Observe, sir, that this resolution holds the tariff of 1828, and every
other tariff, designed to promote one branch of industry at the
expense of another, to be such a dangerous, palpable, and deliberate
usurpation of power, as calls upon the states, in their sovereign
capacity, to interfere, by their own power. This denunciation, Mr.
President, you will please to observe, includes our old tariff of 1816,
as well as all others; because that was established to promote the
interest of the manufacturers of cotton, to the manifest and admitted
injury of the Calcutta cotton trade. Observe, again, that all the
qualifications are here rehearsed, and charged upon the tariff, which
are necessary to bring the case within the gentleman’s proposition.
The tariff is a usurpation; it is a dangerous usurpation; it is a
palpable usurpation; it is a deliberate usurpation. It is such a
usurpation as calls upon the states to exercise their right of
interference. Here is a case, then, within the gentleman’s principles,
and all his qualifications of his principles. It is a case for action. The
constitution is plainly, dangerously, palpably, and deliberately
violated; and the states must interpose their own authority to arrest
the law. Let us suppose the state of South Carolina to express this
same opinion, by the voice of her legislature. That would be very
imposing; but what then? Is the voice of one state conclusive? It so
happens that, at the very moment when South Carolina resolves that
the tariff laws are unconstitutional, Pennsylvania and Kentucky
resolve exactly the reverse. They hold those laws to be both highly
proper and strictly constitutional. And now, sir, how does the
honorable member propose to deal with this case? How does he get
out of this difficulty, upon any principle of his? His construction gets
us into it; how does he propose to get us out?
In Carolina the tariff is a palpable, deliberate usurpation; Carolina,
therefore, may nullify it, and refuse to pay the duties. In
Pennsylvania, it is both clearly constitutional and highly expedient;
and there the duties are to be paid. And yet we live under a
government of uniform laws, and under a constitution, too, which
contains an express provision, as it happens, that all duties shall be
equal in all the states! Does not this approach absurdity?
If there be no power to settle such questions, independent of either
of the states, is not the whole Union a rope of sand? Are we not
thrown back again precisely upon the old confederation?
It is too plain to be argued. Four and twenty interpreters of
constitutional law, each with a power to decide for itself, and none
with authority to bind anybody else, and this constitutional law the
only bond of their union! What is such a state of things but a mere
connection during pleasure, or, to use the phraseology of the times,
during feeling? And that feeling, too, not the feeling of the people
who established the constitution, but the feeling of the state
governments.
In another of the South Carolina addresses, having premised that
the crisis requires “all the concentrated energy of passion,” an
attitude of open resistance to the laws of the Union is advised. Open
resistance to the laws, then, is the constitutional remedy, the
conservative power of the state, which the South Carolina doctrines
teach for the redress of political evils, real or imaginary. And its
authors further say that, appealing with confidence to the
constitution itself to justify their opinions, they cannot consent to try
their accuracy by the courts of justice. In one sense, indeed, sir, this
is assuming an attitude of open resistance in favor of liberty. But
what sort of liberty? The liberty of establishing their own opinions, in
defiance of the opinions of all others; the liberty of judging and of
deciding exclusively themselves, in a matter in which others have as
much right to judge and decide as they; the liberty of placing their
opinions above the judgment of all others, above the laws, and above
the constitution. This is their liberty, and this is the fair result of the
proposition contended for by the honorable gentleman. Or it may be
more properly said, it is identical with it, rather than a result from it.
In the same publication we find the following: “Previously to our
revolution, when the arm of oppression was stretched over New
England, where did our northern brethren meet with a braver
sympathy than that which sprung from the bosom of Carolinians?
We had no extortion, no oppression, no collision with the king’s
ministers, no navigation interest springing up, in envious rivalry of
England.”
This seems extraordinary language. South Carolina no collision
with the king’s ministers in 1775! no extortion! no oppression! But,
sir, it is also most significant language. Does any man doubt the
purpose for which it was penned? Can any one fail to see that it was
designed to raise in the reader’s mind the question, whether, at this
time,—that is to say, in 1828,—South Carolina has any collision with
the king’s ministers, any oppression, or extortion, to fear from
England? whether, in short, England is not as naturally the friend of
South Carolina as New England, with her navigation interests
springing up in envious rivalry of England?
Is it not strange, sir, that an intelligent man in South Carolina, in
1828, should thus labor to prove, that in 1775, there was no hostility,
no cause of war, between South Carolina and England? that she had
no occasion, in reference to her own interest, or from regard to her
own welfare, to take up arms in the revolutionary contest? Can any
one account for the expression of such strange sentiments, and their
circulation through the state, otherwise than by supposing the object