Lec 4. DNA Replication & Repair Flashcards
Which of the three DNA replication complications are Eukaryote specific problems?
Heterochromatin (histones) and Linear chromosomes
What are the three problems encountered during DNA replication?
Heterochromatin (histones), linear chromosomes and incorrect nucleotide added
For Higher Fidelity Repairs what repair mechanisms are used?
Base Excision Repair (BER) and Nucleotide Excision Repair(NER)
For Lower Fidelity Solutions what repair mechanisms are used?
Translesion Synthesis (TLS) and Nonhomologous End Joining (NHEJ)
Why is there a problem with Heterochromatin during DNA replication?
The histones have to be replaced, so theres a mix of new and old histones.
What do recycled histones do?
They retain their marks and thus tell the new histones what marks are needed.
What does the FACT complex do?
Facilitates chromosome transcription by associating with the CMG helicase to displace histones in front of the replication fork
What do chaperone proteins do?
Help histones find their new homes
What is ASF1 stand for?
Anti-silencing function protein 1
What does ASF1 do?
binds H3-H4 dimers and delivers them to the CAF1 complex
What does the CAF1 complex do?
associates with PCNA
Besides during replication what other times do histones need to be remodeled?
Repair, transcription and protein turnover
Why is linear DNA a problem during DNA replication?
DNA polymerase cant fill in the gaps at the ends of chromosomes
What does Telomerase do?
Deals with the linear DNA problem by filling gaps and preventing the shortening of chromosomes
How does Telomerase prevent shortening of chromosomes?
Telomerase is a reverse transcriptase, uses RNA to template DNA. The ssDNA end of the chromosome can invade the upstream telomere sequence.
Which polymerases have an exonuclease domain?
α, δ, ε
What does an exonuclease domain allow?
For proofreading capabilities
Which pol is the best proofreader?
δ
Which pol is the worst proofreader?
α
True or False? Proofreading means that both strands of DNA are synthesized in a discontinous fashion
True
What are the proofreading steps?
Sense incorrect shape, Stall replication fork, Move the nucleotide to exonuclease domain, cleave phosphodiester bond, move growing strand back to polymerase domain.
What mutations are caused by changes in hydrogen bonding potential?
Unfavored Tautomers and modified nitrogenous bases
How are mutations caused by changes in hydrogen bonding potential corrected?
By proofreading and base excision repair
What mutations result from error-prone repair of roadblocks?
Depurination, Covalent adducts with chemicals or between nitrogenous bases
How are mutations that result from error-prone repair of roadblocks corrected?
Translesion synthesis and nucleotide excision repair
How does Base Excision Repair work?
Glycosylases recognize modified nucleotides, nitrogenous base is swung out and clipped off at the glycosidic bond. AP Endonuclease cleaves the phosphodiester bond on either side of the abasic site, single stranded break is created. Pol β or Pol λ add a complementary nucleotide. DNA ligase reforms phosphodiester bonds
How does nucleotide excision repair work?
UV repair enzymes recognize distortion of helix, cleaves some nt surrounding the lesion on single strand. TFII is the helicase that unwinds the region to be cleaved. Pol ε ( replicating cells) or Pol δ (quiescent cells) add back to the complementary nucleotides. DNA ligase reforms phosphodiester bonds.
How does Translesion Synthesis work?
Translesion polymerases take over DNA synthesis at stalled replication forks, each accommodates a particular structure type, does not remove damage, returns control to regular replicative polymerases once lesion is passed. Inserters pick a base to add, extenders continue synthesis.
During Translesion synthesis which are inserters?
Rev1, Pol η, Pol lota
During Translesion synthesis which are extenders?
Pol η, Pol ζ
During Translesion Synthesis what does Rev1 insert?
Can only insert dC, at abasic sites
During Translesion Synthesis what does Pol η insert?
Can insert bases opposite TT dimer (UV rad)
During Translesion Synthesis what does Pol lota insert?
Insert bases opposite snyA and 8-oxoG (ROX)
How are double stranded breaks repaired?
Nonhomologous End Joining and Homologous Recombination
What is Nonhomologous End Joining?
Basically a non-discriminate cut-and-paste event.
How does Nonhomologous End Joining work?
Ends must be prepared for ligation. Double stranded break is recognized by Ku heterodimers, Artemis nuclease cuts at ssDNA-dsDNA junction, TdT, Pol μ and Pol λ bridge the gap, DNA ligase IV adds the final phosphodiester bond
What does TdT do?
Performs untemplated synthesis
