last class 1. transcription 2. rna modification and splicing
DESCRIPTION
Quality control of translation in bacteria Rescue the incomplete mRNA process and add labels for proteasesTRANSCRIPT
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• Last Class
• 1. Transcription• 2. RNA Modification and Splicing• 3. RNA transportation• 4. Translation
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Quality control of translation in bacteria
Rescue the incomplete mRNA process and add
labels for proteases
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Folding of the proteinsIs required before functional
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Folding process starts at ribosome
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Protein Folding PathwayMolecular Chaperone
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An example of molecular chaperone functionsHsp70, early binding to proteins after synthesis
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An example of molecular chaperone functions (chaperonin)Hsp60-like protein, late
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The Fate of Proteins after translation
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E1: ubiquitin activating enzyme; E2/3: ubiquitin ligase
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The production of proteins
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Summary
• RNA translation (Protein synthesis), tRNA, ribosome, start codon, stop codon
• Protein folding, molecular chaperones• Proteasomes, ubiquitin, ubiqutin ligase
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• Control of Gene Expression
• 1. DNA-Protein Interaction• 2. Transcription Regulation• 3. Post-transcriptional Regulation
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Neuron and lymphocyteDifferent morphology, same genome
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Six Steps at which eucaryotic gene expression are controlled
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Double helix Structure
Regulation at DNA levels
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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
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DNA recognition code
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One typical contact of Protein and DNA interfaceIn general, many of them
will form between a protein and a DNA
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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 approach3. Biotechnology to identify protein and DNA
sequence interacting each other.
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Helix-turn-HelixC-terminal binds to major groove, N-terminal helps to position the complex, discovered in
Bacteria
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Homeodomain Protein in Drosophila utilizing helix-turn-helix motif
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Zinc Finger MotifsUtilizing a zinc in the center
An alpha helix and two beta sheet
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An Example protein (a mouse DNA regulatory protein)
utilizing Zinc Finger Motif
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Three Zinc Finger Motifs forming the recognition site
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A dimer of the zinc finger domain of the glucocorticoid receptor (belonging to intracellular receptor family) bound to its specific DNA
sequenceZinc atoms stabilizing DNA-binding Helix and dimerization interface
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Beta sheets can also recognize DNA sequence(bacterial met repressor binding to s-adenosyl methionine)
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Leucine Zipper DimerSame motif mediating both DNA binding and Protein
dimerization(yeast Gcn4 protein)
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Homodimers and heterodimers can recognize different patterns
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Helix-loop-Helix (HLH) Motif and its dimer
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Truncation of HLH tail (DNA binding domain) inhibits binding
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Six Zinc Finger motifs and their interaction with DNA
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Gel-mobility shift assayCan identify the sizes of
proteins associated with the desired DNA fragment
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DNA affinity ChromatographyAfter obtain the protein, run mass spec, identify aa sequence, check
genome, find gene sequence
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Assay to determine the gene sequence recognized by a
specific protein
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Chromatin ImmunoprecipitationIn vivo genes bound to a known protein
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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)
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Gene Expression RegulationTranscription
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Tryptophan Gene Regulation (Negative control)Operon: genes adjacent to each other and are transcribed from a single promoter
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Different Mechanisms of Gene Regulation
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The binding site of Lambda
Repressor determines its
function
Act as both activator and
repressor
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Combinatory Regulation of Lac OperonCAP: catabolite activator protein; breakdown of lactose when glucose is low and lactose is present
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The difference of Regulatory system in eucaryotes and
bacteria 1. Enhancers from far distance over
promoter regions2. Transcription factors3. Chromatin structure
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Gene Activation at a distance
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Regulation of an eucaryotic geneTFs are similar, gene regulatory
proteins could be very different for different gene regulations
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Functional Domain of
gene activation
protein
1. Activation domain and
2. DNA binding domain
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Gene Activation by the
recruitment of RNA polymerase
II holoenzyme
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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
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Gene regulatory proteins help the recruitment and assembly of transcription
machinery(General model)
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Gene activator proteins recruitChromatin modulation proteins to induce transcription
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Two mechanisms of histone
acetylation in gene regulationa. Histone acetylation
further attract activator proteins
b. Histone acetylation directly attract TFs
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Synergistic RegulationTranscription synergy
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5 major ways of gene
repressor protein to be functional
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Protein Assembled to form commplex to Regulate Gene Expression
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Integration for Gene Regulation
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Regulation of Gene Activation Proteins
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Insulator Elements (boundary elements) help to coordinate the regulation
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Gene regulatory proteins can affect transcription process at different steps
The order of process may be different for different genes
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Summary• Gene activation or repression proteins
• DNA as a spacer and distant regulation
• Chromatin modulation, TF assembly, polymerase recruitment
• combinatory regulations