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Last Class Summary
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DNA repair
DNA general recombination
DNA site-specific recombination
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RNA Processing
Polymerase, Promoter, enhancer,
transcription factor, transcription activator
5’ capping, splicing, 3’ cleavage and
polyadenylation
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• From RNA to Protein
The Genetic Code
The Reading Frames
tRNA
tRNA and mRNA pairing
The chemically modified tRNA
Amino Acid Activation
Structure View
Two Steps
Hydrolytic Editing
tRNA synthetases
Hydrolytic Editing
DNA polymerase
Protein synthesis
Ribosome
On endoplasmic reticulum, or free
Ribosome binding sites
Translation:
1. Position at A
2. Peptidyl transferase to transfer
peptide to tRNA at A site
3. Conformational change of large
unit and mRNA on small unit.
Elongation Factor
enhances accuracy
Ribosome
Ribosome
Reaction Mechanism for Protein Synthesis
The Initiation Phase of
protein synthesis in
eucaryotes
Eucaryotic initiation factors
eIFs
Bacterial one mRNA several gene
Eucaryotes one mRNA, one gene.
Stop codon
Human tranlation release factor and tRNA
Multiple Copies on the
Same mRNA
Rescue the incomplete
mRNA process and add
labels for proteases
Incorporation of selenocysteine into a growing polypeptide chain
Folding of the proteins
Molten Globule
Folding process starts at ribosome
Protein Folding Pathway
Molecular Chaperone
An example of molecular chaperone functions
Hsp70, early
An example of molecular chaperone functions
Hsp60-like protein, late
The Fate of Proteins after translation
Proteasome
Ubiquitin
E1: ubiquitin activating enzyme; E2/3: ubiquitin ligase
Protein aggregation and human disease
Cross-beta filaments
The production of proteins
Summary
• RNA translation, tRNA, ribosome, start
codon, stop codon
• Protein folding, molecular chaperones
• Proteasomes, ubiquitin, ubiqutin ligase
• Protein aggregation
• Control of Gene
Expression
• DNA-Protein Interaction
Neuron and lymphocyte
Experiment to show that the differentiated cell contains al the
genetic information
Differences in mRNA
expression in different
cancer cells
Differences in Protein expression in different cancer cells
Red: common; Blue: specific
Six Steps at which eucaryotic gene expression are controlled
Double helix Structure
The outer surface difference of base pairs without
opening the double helix
DNA recognition code
DNA bending induced by protein binding
Helix-turn-Helix
Proteins contain Helix-turn-Helix
Homeodomain Protein is a special
class utilizing helix-turn-helix
Zinc Finger Motifs
An Example protein
utilizing Zinc Finger Motif
B-sheets can also recognize DNA
Leucine Zipper Dimer
Homodimers and heterodimers can
recognize different patterns
Covalently-joint dimer
Helix-loop-Helix dimer
Single truncation of HLH inhibits binding
One of the most Common
protein-DNA interaction
Six Zinc Finger motifs and their interaction with DNA
Gel-mobility shift assay
DNA affinity Chromatography
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 or proteins bound
to known sequences