171030 DNA Replication and Repair-3.pdf

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1、Content•DNAdamage•TheDNAdamageresponse•DNArepair–Directreversal–Baseexcisionrepair–NucleotideexcisionrepairPaulModrichreceivingNobelprize–Translesionsynthesis–Mismatchrepair–Doublestrandbreakrepair•Recombinationrepair•Non-homologousendjoiningWhatisDNAMismatch?Whatismismatch?Non-Watso

2、n-CrickbasepairsTwoclassesofmismatches•Base-basemismatchG-T,A-C,A-A,A-G,G-G,C-C,C-T,T-T•Insertion-deletionmismatchCACACACACACACACACACACACACACACACAGTGTGTGTGTGTGT-GTGTGTGTGTGTGTCACellularmechanismsremovingmismatchesGT5’3’DNApolymeraseproofreadingDNAmismatchrepair(MMR)exonucleaseGG5’T5’

3、3’3’G5’ResumingDNA3’synthesisGG5’CC3’5’3’MismatchRepairEnsuresReplicationFidelityAAAA-AAAAParentalstrandTTTTTTTTDaughterstrand5’TT3’TDaughterstrandGParentalstrandMismatchrecognition&excisionAAAAAAAA5’3’MismatchrepairisGtargetedonlytotheRepairresynthesisnewlysynthesizedstrandAAAAAAAAT

4、TTTTTTT5’3’CGKeymismatchrepairComponentsE.coliHumanFunction/activityMutSMutSα(MSH2-MSH6)Base-basemispairrecognitionMutSβ(MSH2-MSH3)Insert./del.MispairrecognitionMutLMutLα(MLH1-PMS2)Endonuclease(?)MutLβ(MLH1-PMS1)?MutLϒ(MLH1-MLH3)?MutH?EndonucleaseUvrD?HelicaseExoI,ExoVII,ExoIExonucle

5、aseExoX,RecJSSBRPAssDNAbindingPolIIIPolδDNAsynthesisDNAligaseLigaseINickligationRegulationofreplicationinitiationbymethylation•ReplicationofmethylatedDNAgiveshemimethylatedDNA,whichmaintainsitsstateatGATCsitesuntiltheDammethylaserestoresthefullymethylatedcondition.•Replicationgenerat

6、eshemimethylatedDNA,whichcannotinitiatereplication.•Thereis~a10mindelaybeforetherepeatsareremethylated.Howismismatchrepairtargetedtothenewlysynthesizedstrand?Methyl-DirectedMismatchRepairinE.coliCH3G-A-T-CC-T-A-GMutH(endonuclease)CH3G-A-T-CC-T-AGHowismismatchrepairtargetedtothenewlys

7、ynthesizedstrand?Nick-DirectedMismatchRepairinEukaryoticcells5’G3’TMutLMutSPCNAEXO1AAAA-AAAAParentalstrandTTTTTTTTDaughterstrand5’5’3’TT3’TDaughterstrandGParentalstrandRPA5’3’Pold,PCNARFC,ligaseI5’G3’C•EukaryoticcellsrelyonexonucleaseI(Exo1)forMMR•Exo1onlycontainsa5’to3’exonuclease

8、activityHowi