If you place the fully protonated form

of lysine in aqueous solution at pH

7.0, what will happen to the pH?

A. The pH will increase.

B. The pH will decrease.
C. The pH will not change.
D. It is impossible to predict what change will
happen.
 Introductory
Biochemistry
Protein Structure and Function
Peptide Bonds & Primary Structure
Formation of a Peptide Bond

amide bond
Peptide Nomenclature I
 Two amino acids joined by one peptide form
a “dipeptide”

 Similarly:
 Three amino acids – tripeptide
 Four amino acids – tetrapeptide
 Five amino acids – pentapeptide
 etc.
Peptide Nomenclature II
 Peptides/oligopeptides
 General term for a larger number of amino acids,
often refers to synthetic peptides (< 40 residues)

 Polypeptide
 Long chain of amino acids, usually produced naturally

 Protein
 Large polypeptide (or >1 polypeptide) with a biological
function
A dipeptide (containing Met and Arg).

Arg-Met Met-Arg

From: https://pepdraw.com/
A Tetrapeptide

Only the terminal amino and carboxylate groups in a peptide

retain their charge. The others are eliminated by the formation of
peptide bonds.
Side chains retain their charge (if they have one).
 The following molecule is:

A. A tetrapeptide with 5 peptide bonds.

B. A pentapeptide with 5 peptide bonds.
C. A pentapeptide with 6 peptide bonds.
D. A hexapeptide with 5 peptide bonds.
How many charged functional
groups are present on the peptide
below at physiological pH (7.0)?

A. 1
Arg-Asp-Cys-Tyr-Gln-Val-Glu
B. 2
C. 3
D. 4
E. 5
Primary Structure
 The sequence of amino acids in a
polypeptide is the “primary structure”
 Covalent peptide bonds join each amino acid to
the next
 Every protein or polypeptide has a unique
sequence

 Peptide bonds have distinguishing chemical

characteristics
 Rigid
 Planar
 Properties of Peptide Bonds
 The electrons in peptide
bonds are somewhat
delocalized generating two
resonance forms.

 Peptide bonds therefore

exhibit partial double-bond
character with no rotation
around the C-N bond.

 The functional groups in

peptide bonds are potential
H-bond acceptors or donors.
Peptide Bonds are Rigid & Planar

Rotation around the C-C and N-C bonds can

occur to a certain degree

The backbone of a polypeptide includes the

C atoms and those involved in the peptide bond
How many charged side chains are
present on the peptide below at
physiological pH (pH 7.0)?

A. 1
B. 2 Asn-Trp-Cys-Tyr-Lys-Val-Glu
C. 3
D. 4
E. 5
 Introductory
Biochemistry
Protein Structure and Function
Secondary Structure
Levels of Protein Structure

Primary structure determines the 3-D structure

3-D structure determines function

The Rotation of Polypeptide
Backbones is Limited
folding conformations are limited
H-Bonds in Polypeptide
Backbones
The chemical groups found in
peptide binds are highly polar.
d-
Carbonyl groups are hydrogen
d+ bond acceptors, NH groups are
hydrogen bond donors.
d-
d+
Secondary Structure
 Local folding of the polypeptide backbone
 Allows for hydrogen bonding of the groups in
the polypeptide backbone (C=O, N‒H)
 “Regular” secondary structures occur when
every amino acid in a segment of the
polypeptide adopts the same geometry.
 A few regular patterns occur:
 The a-helix
 The b-sheet
 An a-Helix
The carbonyl oxygen
of each residue forms
a H-bond with the
backbone –NH group
four residues downstream
C1 N5
C2N6

Except for amino acid residues at

either end, all the backbone CO
and NH groups are hydrogen
bonded to one another in the helix
What is the net charge of the peptide
below at physiological pH (~7)?

Ala-Arg-Asn-Lys-Glu-Ser-Gly
A. -2
B. -1
C. 0
D. +1
E. +2
Source: http://bioinfo.au-kbc.org.in/books/bi/bi.html
Side Chains in an a-Helix

Helix is SOLID with atoms in

the polypeptide backbone in
van der Waals contact with one
another in the center.

Amino acids project outwards,

residues 3-4 apart in the
primary structure are close in
the secondary structure.

-helix same diameter as major groove

The following sequence of amino
acids is very unlikely to fold into a
stable a-helix at pH 7.0. Why?

Asp-Glu-Ser-Asp-Glu-Glu-Arg-Val-Asp-Ala-Glu-Asp

A. Steric hindrance among side chains.

B. Lack of appropriate groups for H
bonding among side chains.
C. Electrostatic repulsion among side
chains.
D. None of the above.
Why is proline not common in the middle of an -helix?
Parallel and Anti-Parallel b-Sheets
Multiple β-strands are arranged side by side.
Strands are joined by loops or other structures.
b-sheets are often drawn as arrows

The arrows are shown pointing from the N-terminal end to the C-terminal end.
The example on the left shows an anti-parallel sheet, the one on the right a parallel
sheet. Irregular structures connect the strands in these examples.
An anti-parallel β-sheet
Side view of two anti-parallel
strands in a b-sheet

Side chains are located above and below the

plane of the sheet
What forces stabilize -helices
and -sheets?
 -helices: H-bonds between backbone CO
and NH groups in the same helices

 -sheets: H-bonds between backbone CO

and NH groups of neighbouring strands
Proteins contain regular and irregular
secondary structures.
 -helices and -sheets are regular secondary
structures. The peptide backbone has the same
configuration/conformation for every amino acid
within the secondary structure.

 Distinct elements of regular secondary structure are

linked together by polypeptide loops of various sizes
ranging from simple hairpins to longer loops. These
structures are irregular.
Irregular Secondary Structure
Irregular does NOT
mean disordered!

Polypeptide loops typically

have a backbone of
irregular secondary
structure.
Which of the following statements about
-helices and -sheets is/are FALSE?

A. Both are stabilized by hydrogen bonds between groups

in the polypeptide backbone.
B. Both have conformations that require specific angles of
rotation around the peptide (C-N) bond.
C. Both -helices and -sheets form only from consecutive
amino acid residues in the polypeptide.
D. A and B
E. B and C
 Introductory
Biochemistry
Protein Structure and Function
Tertiary Structure & Protein
Folding
 Tertiary Structure
 Arrangement of all atoms in a single polypeptide
 Arrangement of secondary structures in relation to
one another
 Positions of amino acid sidechains
 Prosthetic groups (heme, FAD, etc.)

 Can be classified into two general morphologies:

 Fibrous (elongated)
 Globular (compact)
Two main protein classes
 Fibrous proteins
 practically insoluble in aqueous solutions
 form long protein filaments-limited residues with repeats
 usually structural or connective proteins
 Globular proteins
 basically soluble in aqueous solutions
 fold into compact structures with nonpolar cores and
polar surfaces
 Fibrous Proteins

Fibrous proteins (like collagen,

shown here), tend to adopt linear,
extended structures.
Some globular proteins (at the same scale)

Myoglobin

Lysozyme

Hemoglobin

Chymotrypsin
Hexokinase
Lactate dehydrogenase Space-filling diagrams
Tertiary structure in globular proteins is
highly variable
Myoglobin

Lysozyme

Hemoglobin

Chymotrypsin
Hexokinase
Lactate dehydrogenase Ribbon diagrams
 Hydrophobic interactions and protein 3-D
shape in a soluble globular protein

Hydrophobic side chains are Hydrophilic side chains are

most likely to be found in the most likely to be found on the
interior of a globular protein surface of a globular protein

In these side-by-side figures of a protein, the hydrophobic side chains are indicated
in the left image, and the polar and charged side chains in the right image. The
backbone here is simplified as a ribbon.
 Regular secondary structure found in
the interior of folded proteins

Loops tend
to be located
on the surface

Hydrophilic residues

Hydrophobic residues

“Copyright by Pearson; used with permission”

a-helix and b-sheet secondary structure is
abundant/enriched in the interior of folded
proteins, whereas irregular loops occur on
the outside. Why?

A. The loops are needed for the polypeptide to “turn corners”

and this occurs on the outside of the protein.
B. a-helix and b-sheet structures are more compact than
loops, and so they fit better in the interior of the folded
protein.
C. The side chains in a-helix and b-sheet structures are mostly
non-polar.
D. The H bonding requirements of the polypeptide backbone in
irregular loops are not fully satisfied, and so these
structures interact with water at the surface.
The Hydrophobic Effect in
Protein Folding
The shape of globular
proteins depends on the
relative positions of
hydrophobic amino acids
in the protein’s 1°
structure.

The hydrophobic effect is

the “driving force” via
which soluble globular
proteins adopt and
maintain their 3°
structure.
Hydrophobic side chains
Polar side chains
Hydrogen bonding
 Weak forces between closely-positioned polar side
chains. These help to “fine tune” and stabilize 2° and 3°
structures.

Hydrogen bonds can also

form between backbone
groups and side chains.
Which of the following amino acid residues can
form hydrogen bonds with Ser residues located in
the middle of an -helix?

A. Residues in a neighbouring -helix.

B. Residues located within the same -helix.
C. Both A and B.
D. Neither A nor B.
Ion Pairs (“Salt Bridges”)
 Electrostatic interactions between closely-positioned
formal charged groups. Like H-bonds, these help to “fine
tune” and stabilize 2° and 3° structures.

Can form between positively and

negatively charged groups

+ charges - charges

N-terminus C-terminus
Lys, Arg Asp, Glu
(His) (Tyr, Cys)

Charges will depend on pH of environment

Disulfide bonds/bridges
 Covalent bonds between closely-positioned cysteines.
 These form stabilizing crosslinks for extracellular
proteins (or proteins in the lumen).

Disulphide bonds in
Ribonuclease A, an
extracellular protein.

In the cytosol, cysteines

do not oxidize to cystine
as it is a reducing
environment.
Protein folding
Protein folding
 Which of the following statements is FALSE
with respect to the ribbon diagrams shown
below ?
(1) (2) (3)

A. Protein 2 contains anti-parallel b-sheet strands.

B. Protein 1 contains parallel b-sheet strands.
C. The secondary structure of protein 3 is all a-helix.
D. These proteins have both secondary and tertiary structure.
Protein structure
 Domain
 A polypeptide segment that has folded into a
single structural unit with a hydrophobic core.
 Proteins may contain more than one domain.

 Motif
 A short region of polypeptide with a recognizable
3D shape.
 Zinc fingers (example)
 May be found in many contexts.
Which of the following amino acid residues can
form hydrogen bonds with Ala residues located in
the middle of an -helix?

A. Residues in a neighbouring -helix.

B. Residues located within the same -helix.
C. Both A and B.
D. Neither A nor B, because Ala is unable to form
hydrogen bonds.
Protein Domains
Pyruvate kinase (left) is composed of a
single polypeptide chain, which can be
divided into 3 domains.

Cow gamma crystalline (below) contains

two domains in one polypeptide.
Motifs in Protein Structure
These patterns/arrangements
of secondary structures are
examples of different
structural motifs commonly
found in proteins.
Zinc Fingers: An example of a structural
motif including a prosthetic group.

(a) A single Zn2+ ion coordinated by

2 Cys and 2 His residues.

(b) Two Zn2+ ions coordinated by 6

Cys residues.
 Prosthetic groups
 Some proteins have prosthetic groups as part of
their tertiary structure.
 Chemical reactivity of amino acid side chains is limited

 Prosthetic group: A non-peptide component that is

permanently incorporated into a protein. These
provide:
 Structure – e.g. Zn2+ in zinc fingers
 Functional chemical groups – e.g. Heme in hemoglobin
Heme is a Prosthetic Group
Globular proteins are stabilized by weak
noncovalent forces and are easily unfolded or
“denatured”.

 Heat H-bonds/hydrophobic interactions

 Changes in pH salt bridges/H bonds
 Salt salt bridges/ion pairs
 Detergents hydrophobic interactions
 Breaking Disulphide Bonds

Cys Cys Cys

Cys Cys Cys

 Reducing agents (DTT) can disrupt disulphide bridges

Quaternary structure
Proteins composed of more than one polypeptide chain
Each polypeptide chain is called a subunit
Quaternary structure
 Named by number and type of subunits
 2 subunits: Dimer
 Identical subunits: Homodimer
 Non-identical subunits: Heterodimer
 3 Subunits: Trimer
 Tetramer, pentamer, etc.

 Stabilized by the same forces as 3° structure

 Hydrophobic interactions
 H-bonds, ion pairs “fine-tune”
Hemoglobin contains 2 a subunits and 2 b
subunits. How could we describe this
protein’s quaternary structure?

A. A homodimer.
B. A heterodimer.
C. A homotetramer.
D. A heterotetramer.
E. None of these.
Which of the following statements
about quaternary structure is TRUE?
A. Quaternary structure is defined as the arrangement of
polypeptide backbones in proteins with four subunits.
B. Quaternary structure exists only in monomeric proteins
containing more than one domain.
C. Quaternary structure is stabilized by the same types of
noncovalent interactions as tertiary structure.
D. Quaternary structure requires covalent interactions
between polypeptide chains.
E. C and D are both true.
Polar amino acids in the hydrophobic core
of a globular protein are involved in either
H-bonding or ion pairs.

A. True
B. False