Courses/Bioc 406
Quiz: How does E. Coli discriminate between template strand and newly synthesized strand? (Mismatches caused by error in DNA replication are corrected by a repair system that must discriminate between template strand and newly synthesized strand) MutH binds to the methyl group of the old strand, and endonucleases thew new strand when it reaches MutL and MutS complex. The new strand is not methylates (that's how MutH knows which is which) Which is true about base-excision DNA repair? It uses DNA glycosylases It uses DNA helicases It removes methyl groups from methylated DNA bases IT cleaves the damaged DNA on either site of the lesion
What is the role of exinucleases in nucleotide-excision repair of DNA? The cleave the damaged DNA strand at either side of a lesion DNA damage caused by sunlight is most commonly repaired by: Nucleotide-excision repair, and DNA photolyase (only bacteria) Deamination of cytosine leads to formation of: uracil (this leads to U-G matching BAD) Pol I synthesizes new DNA with very high fidelity due to its: nascent 3'-5' exonuclease in proofreading site (palm) Mutation impairing 3'-5' exonuclease activity on DNA polymerase faster and less accurate DNA replication Which of the following sequences is most likely to have indel mutations? GTGTGTGTGTG. 2,4,6 base pair insertion/deletions because we have it in units of 2
Three flavors of mutation reduction Inherent accuracy error rate: low error rate proofreading: lower error rate surveillance and repair: really low error rate (evolutionary context: these mechanisms drive [[mutation|mutation rate]] and [[selection|mutation-selection balance]]) [[Tautomer]] shifting electrons around tautomer of adenine hydrogen bond differently, bind to cytosine instead of thymine all 4 bases tautomers [[DNA polymerase]] proofreads the nascent strand the 'palm' nascent 3'-5' exonuclease in proofreading site independent of active site checking the DNA (DNA Pol 1 has TWO exonuclease activities) Chemical mutagens Alkylating agents: nitrogen mustard, ethylnitrosourea, MNNG methylguanine base pairs with either T or C MGMT: suicide enzyme, prevents G:C to A:T mutations Oxidizers cytosine->uracil adenine->hypoxanthine Fix: Base excision repair 1. defective base removed by a DNA glycosylase. several glycosylases recognize different problem bases 2. backbone is cleaved at the ATP site by AP endonuclease 3. DNA polymerase uses its dRP lyase activity to remove the naked sugar/phosphate and fills in gap, then sealed by DNA ligase Uracil-DNA glycolyase catalyzes base excision base flips the dU:dG mismatch, cleaves Why thymine: excision repair knows that when dU is present, it is almost certainly the product of deamination of dC! Why Uracil at all: DNA intercalating agents cause indels repetitive DNA sequences are susceptible to slipped-strand mispairing, which can cause indels can slip forward (+1 amino acid insertion), slip backward (-1 amino acid deletion) Huntington's: repeat of >40 caused by this slippage^
Cancer — somatic cell evolution driven by [[selection]] (cells with proliferative mutations outcompete normal cells) mutations in protooncogenes: High cell division dominant Antioncogenes: Low inhibitor of cell division -> high cell division recessive DNA damage base loss: 20,000/cell/day deamination: 100/cell/day 1/1000 of the genome is damaged per year Radiation carbons one sugar phosphate ring bind to carbons on adjacent ring Fix: DNA photolyase (not in humans) 1. MTHF absorbs UV-A, gets photoexcited 2. transfers excitement to FADH 3. FADH transfers electron to dimer, breaking, into thymine monomers, takes electron back Fix: Nucleotide excision repair UvrABC endonuclease (ABC exinucleases) recognizes backbone distortions helicase comes in and removes distorted segment Pol 1, DNA ligase fixes gap [[Xeroderma pigmentosum]] genetic disease of excision repair DNA mismatch repair repairing mistakes during replication 1. MutS recognizes mismatch 2. MutL recognizes mismatch 3. Use Atp. MutS and MutL pulls DNA towards itself, creating a loop away from the rest of the DNA 4. mutH endonuclease recognizes methyl group on DNA (methyl group is on GATC palindrome sequence), runs into MutL-MutH complex 5. MutH has endonuclease activity and cleaves just the new strand 6. UvrD and exnonuclease helicase unwind the two strands 7. Fills in with Pol I and ligase