Specific DNA repair Flashcards
2 sources of mutation
I. mistakes during DNA replication (occasional errors)
II. Chemical/environmental factors
3 types of chemical/environmental sources of mutations
-heat -> deamination
-chemicals -> add groups to bases or sugars, break bonds, fuse parts
-irradiation -> DNA absorbs energy of photons, leads to breakage or fusion of molecular parts
Survival Curves: exposure to radiation/chemicals leads to ______. survival curves indicate _______ vs _______
-inability to form colonies or plaques
- fraction of surviving population (colonies, plaques)
- exposure/dose
Evidence that chemicals/irradiation kill by damaging DNA: UV irradiation -> DNA absorbs light at ____; protein at ____. UV light at ___ much more effective at killing cells so ___ is target molecule
-260 nm
-280 nm
-260 nm
-DNA
Evidence that chemicals/irradiation kill by damaging DNA: Mutant strains defective in DNA repair are more sensitive to killing by mutagens
i) Block in DNA replication (EX. ______ breaks)
These are mutagens that do not affect base pairing
ex. UV light causes production of _____ dimers
Dimers are still correctly _____ to bases in opposite strand. ______ recognizes distortion in DNA helix caused by dimer, tries to proofread- can never fix problem so protein remains stalled
ii) Double strand breaks - no mechanism to repair double strand breaks -> ______
- ssDNA
- pyrimidine
- base paired
- DNA polymerase
-cell death
Evidence for DNA repair: Survival curves are not ______. initial plateau followed by ______. efficient repair occurs at _____ and bacteria/phage do not die. at _____ doses, repair systems become saturated or damaged leading to a ________
-linear
-downward curve
- small UV doses
-higher
-steep drop in survival
Evidence for DNA repair: Photoreactivation: survival of irradiated bacteria is increased when ______ before plating for colonies. due to activation of a _____ enzyme by ______ that can cleave T-T dimers. can also be observed with ______ that are UV-irradiated and plaqued on bacteria that have been exposed to ______
-cells are exposed to sunlight
- photolyase
-visible light (300-600 nm)
-phages
-visible light
Evidence for DNA repair: DNA Repair Mutants: Mutants have diminished capacity to survive ______. Mutations are in genes encoding ________
- mutagenic treatments
-DNA repair enzymes
2 kinds of repair pathways + def.
-specific repair pathways (recognize specific alterations to the DNA)
-general repair pathways (recognize distortions in DNA caused by a wide range of damage)
5 specific repair pathways
- deaminated bases
- VSP repair
- oxidative repair
- alkylation repair
- pyrimidine dimers
4 general repair pathways
- CH3-directed mismatch repair
- nucleotide excision repair
- DNA damage tolerance mechanisms
- SOS inducible repair
Many specific repair pathways use damage-specific enzymes to perform similar steps: ______ specific for the damage cleaves the base from the sugar. _______ cleaves the DNA backbone on one side of the damaged base (usually 5’). 5’->3’ exonuclease activity of _________ degrades damaged strand. _______ uses free 3’-OH as a primer to replace damaged DNA. ______ seals gap
① glycosylase enzyme
②an AP (apurinic or apyrimidinic) endonuclease
③ DNA POL I (repair polymerase)
④ DNA POL I
⑤ DNA ligase
Repair of Deaminated Bases: specific DNA glycosylases remove ______. AP endonuclease cuts _____. 5’ exonuclease activity of ______ degrades strand ahead of pol. DNA pol I uses _____ as primer to synthesize more DNA.______ seals gap.
-damaged base
-backbone
-DNA pol I
-3’-OH
-DNA ligase
Very Short Patch (VSP) Repair happens for mutations not recognized by _______. ex. occurring after ______. ______ binds TG mismatch = makes break next to T. T removed and resynthesized by ____
_____ = seals gap
very specific; Vsp endonuclease gene is encoded beside ______ which creates ______ = coevolution
-mismatch repair;
- replication / methylation
- VSP endonuclease
-DNA pol I
-DNA ligase
-Dcm methyltransferase gene
-5-CH3-Cystosine
Repair of Oxidative Damage: reactive oxygen forms generated by: ______ . damage DNA, most common lesion is oxidized G = _______. 8-oxoG mispairs with ___; _______ transversions
-metabolism, environment, chemicals
-7,8-dihydro-8- oxoguanine (8-oxoG)
-A
-GC =TA and TA = GC
mut genes: mutD -> ______
mutS, mutL, mutH -> ______
-DNA pol III editing ; dnaQ
-CH3 directed mismatch repair
Bacteria have evolved 3 ways of fixing oxidative damage:
_____ (N-glycosylase/AP endonuclease) -> removes 8-oxoG from DNA
_____ (N-glycosylase) -> removes A incorrectly paired with 8-oxoG
_____ (phosphatase) -> degrades 8-oxodGTP -> 8-oxodGMP
-MutM
-MutY
-MutT
We know that MutM, MutY, MutT function independently of one another because mutations in each are_____
additive
alkylating agents add ____ groups to bases or phosphates.can alter bases leading to ____ which creates distortions of helix. ____ distortions are repaired by the general repair pathways. _____ distortions are repaired by specific repair pathways in 3 different ways:
-alkyl
- mispairing
-major
- subtle
Alkylation repair: Adaptive response – ADA protein.
Brief treatment with alkylating agent allows increased _____ upon subsequent treatments
- exposure to alkylating agents induces _____ that can repair damage
- methylation of N-terminus of ______ converts it to a transcriptional activator (recruits _____)
- induces expression of an _____ proteins that can help fix
alkylation damage
-survival
-expression of genes
-ada methyltransferase
-RNA pol
-adaptive response
Alkylation repair: N-Glycosylases
____ = removes 3-CH3-G, 3-CH3-A, 7-CH3-G
-AlkA
Alkylation repair: ______ remove alkyl groups from O6 of G and O4 of T by transferring to themselves
- inactivated by transfer of _____
- e.g. transfer to ADA protein C-terminus = ______
-Methyltransferases
-alkyl group
-suicide inactivation
alkylation repair enzymes are inducible
exposure to alkylating agents induces expression of genes that can repair damage(eg. _____)
- methylation of _____ of ada methyltransferase converts it to a ______
-induces expression of adaptive response proteins that can help fix _____
-Ada, AlkA
- N-terminus
-transcriptional activator
-alkylation damage
Pyrimidine Dimer Repair: _______ is a major source of natural damage to DNA
Bases strongly absorb _____ -> energy increases reactivity of
double bonds -> formation of abnormal bonds that link bases
Repaired by ______: repairs ______ dimers; first DNA repair system to be discovered
* 1940s, Albert Kelner -> Streptomyces griseus more likely to survive UV
irradiation in ______
*______ carrying FADH2 binds to fused bases
* absorption of visible light (300-600nm) provides energy to allow ______
-UV irradiation
-UV light
-photoreactivation
-cyclobutane & pyrimidine
-the light than in the dark
-photolyase
- separation of fused bases
