• Last Class

• 1. Transcription
• 2. RNA Modification and Splicing
• 3. RNA transportation
• 4. Translation
 Quality control of
translation in bacteria

Rescue the incomplete

mRNA process and add
labels for proteases
 Folding of the proteins
Is required before functional
Folding process starts at ribosome
Protein Folding Pathway
Molecular Chaperone
An example of molecular chaperone functions
Hsp70, early binding to proteins after synthesis
An example of molecular chaperone functions (chaperonin)
Hsp60-like protein, late
The Fate of Proteins after translation
E1: ubiquitin activating enzyme; E2/3: ubiquitin ligase
The production of proteins
 Summary
• RNA translation (Protein synthesis), tRNA,
ribosome, start codon, stop codon
• Protein folding, molecular chaperones
• Proteasomes, ubiquitin, ubiqutin ligase
 • Control of Gene Expression

• 1. DNA-Protein Interaction
• 2. Transcription Regulation
• 3. Post-transcriptional Regulation
 Neuron and lymphocyte
Different morphology, same genome
Six Steps at which eucaryotic gene expression are controlled
 Regulation at DNA
levels

Double helix Structure

 The outer surface difference of base pairs without
opening the double helix

Hydrogen bond
donor: blue
Hydrogen bond
acceptor: red
Hydrogen bond:
pink
Methyl group:
yellow
DNA recognition code
 One typical contact of
Protein and DNA interface
In general, many of them
will form between a
protein and a DNA
 DNA-Protein Interaction

1. Different protein motifs binding to DNA: Helix-

turn-Helix motif; the homeodomain; leucine
zipper; helix-loop-helix; zinc finger
2. Dimerization approach
3. Biotechnology to identify protein and DNA
sequence interacting each other.
 Helix-turn-Helix
C-terminal binds to major groove, N-terminal
helps to position the complex, discovered in
Bacteria
Homeodomain Protein in Drosophila
utilizing helix-turn-helix motif
 Zinc Finger Motifs
Utilizing a zinc in the center
An alpha helix and two beta sheet
An Example protein (a mouse
DNA regulatory protein)
utilizing Zinc Finger Motif
 Three Zinc Finger Motifs
forming the recognition site
 A dimer of the zinc finger domain of the glucocorticoid receptor
(belonging to intracellular receptor family) bound to its specific DNA
sequence
Zinc atoms stabilizing DNA-binding Helix and dimerization interface
 Beta sheets can also recognize DNA sequence
(bacterial met repressor binding to s-adenosyl methionine)
 Leucine Zipper Dimer
Same motif mediating both
DNA binding and Protein
dimerization
(yeast Gcn4 protein)
Homodimers and heterodimers can
recognize different patterns
Helix-loop-Helix (HLH) Motif and its dimer
Truncation of HLH tail (DNA binding domain) inhibits binding
Six Zinc Finger motifs and their interaction with DNA
 Gel-mobility shift assay
Can identify the sizes of
proteins associated with the
desired DNA fragment
 DNA affinity Chromatography
After obtain the protein, run mass spec, identify aa sequence, check
genome, find gene sequence
Assay to determine the gene
sequence recognized by a
specific protein
 Chromatin Immunoprecipitation
In vivo genes bound to a known protein
 Summary
• Helix-turn-Helix, homeodomain, leucine
zipper, helix-loop-helix, zinc-finger motif
• Homodimer and heterodimer
• Techniques to identify gene sequences
bound to a known protein (DNA affinity
chromatography) or proteins bound to
known sequences (gel mobility shift)
Gene Expression Regulation
Transcription
 Tryptophan Gene Regulation (Negative control)
Operon: genes adjacent to each other and are transcribed
from a single promoter
Different Mechanisms
of Gene Regulation
The binding site of
Lambda
Repressor
determines its
function

Act as both
activator and
repressor
 Combinatory Regulation of Lac Operon
catabolite activator protein; breakdown of lactose when glucose is low and lactose is pres
The difference of Regulatory
system in eucaryotes and
bacteria
1. Enhancers from far distance over
promoter regions
2. Transcription factors
3. Chromatin structure
Gene Activation at a distance
 Regulation of an eucaryotic gene
TFs are similar, gene regulatory
proteins could be very different for
different gene regulations
 Functional
Domain of
gene
activation
protein

1. Activation
domain and
2. DNA
binding
domain
Gene Activation
by the
recruitment of
RNA polymerase
II holoenzyme
 Gene engineering revealed the function of
gene activation protein
Directly fuse the mediator protein to
enhancer binding domain, omitting activator
domain, similar enhancement is observed
 Gene regulatory proteins help the
recruitment and assembly of transcription
machinery
(General model)
Gene activator
proteins recruit
Chromatin
modulation proteins
to induce
transcription
Two mechanisms of
histone
acetylation in
gene regulation
a. Histone
acetylation
further attract
activator proteins
b. Histone acetylation
directly attract TFs
Synergistic Regulation
Transcription synergy
5 major ways
of gene
repressor
protein to be
functional
Protein Assembled to form commplex to
Regulate Gene Expression
Integration for Gene Regulation
Regulation of Gene Activation Proteins
Insulator Elements (boundary elements) help to
coordinate the regulation
Gene regulatory
proteins can affect
transcription process
at different steps

The order of process

may be different for
different genes
 Summary
• Gene activation or repression proteins

• DNA as a spacer and distant regulation

• Chromatin modulation, TF assembly,

polymerase recruitment

• combinatory regulations