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Broad categories of DNA repair
- Pre-replicative
- Post-replicative
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Pre-replicative
Scan current template DNA for irregularities
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Post-replicative
Scan nascent DNA for for errors
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Spontaneous errors
- Result from mismatches during replication
- Not detected by proofreading
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Induced errors
Reaction of Mutagen with parent DNA
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Hydrolytic bonds in DNA
- Phosphodiester
- N-glycosyl (depurination)
- Bonds linking amine groups to rings
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Types of DNA repair mechanisms
- Direct
- Excision
- Mismatch
- Nonhomologous end-joining
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Direct repair is used for
Guanine methylation
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Nucleotide excision repair is used for
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Varieties of Nucleotide excision repair
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Mismatch repair is used for
Mismatched bases
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Base excision repair is used for
- Single strand breaks
- Single-base damage
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Homologous repair is used for
Double strand breaks
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Non-homologous end-joining is used for
Double strand breaks
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Why are polymerases so accurate
- Nucleotide selection processes
- 3'-5' proofreading (not all)
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What is it called when polymerase makes a mistake
Misincorporation
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E.coli mismatch protein with endonuclease activity
- MutL
- (If in doubt, this is correct - see notes, March 30th)
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When does a mismatch or misincorporation become a mutation?
When it is replicated
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Polymerase fidelity (alone, w/ proofreading, w/ MMR)
- 1 in 10^5
- 1 in 10^7
- 1 in 10^10
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Sequence recognized in MMR?
GATC
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Frequency of GATC
~1 in 256 bases
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Hemimethylated DNA
Only one methylated strand
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MutS
- Prokaryote MMR
- Dimer
- Scans for mismatches
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Steps in MMR
- MutS binds (ATP produced) and scans the DNA
- If a mismatch if found, mutL is recruited
- MutH is recruited by myL and nicks the DNA
- UvrD metls from the nick to the mismatch
- Exo I (5->3) or Exo VII (3->5) degrade the section
- Pol III fills in the gap
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MutH
- Prokaryote MMR
- Endonuclease
- Nicks nearest unmethylated GATC to mismatch
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UvrD
- Prokaryote MMR
- Helicase (II)
- Melts from the nick to mismatch
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Prokaryote MMR Proteins without eukaryote homologs
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What does PMS stand for?
Post meiotic segregation
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MutSα structure and activity
- Six heterodimers of MSH2 and MSH6
- Bind mismatches or insertions/deletions (small)
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MutSβ structure and activity
- Also six heterodimers
- Only bind to insertion/deletions (all sizes)
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Eukaryote MMR Overview
- MutSα and MutSβ will scan for defects
- Binding of MutSα and MutSβ
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