分子生物學(xué)教學(xué)課件：chapter9 The mutability and repair of DNA

Chapter9:

Chapter9ThemutabilityandrepairofDNACausesRepairMutationRecombinationHomologousrecombination(同源重組）Site-specificrecombination(位點(diǎn)專一重組)

Transposition(轉(zhuǎn)座）ThecausesofmutationMutagensErrorinreplicationchemicalphysicalEnsuringtheaccuracyReplicationerrorscausetypesPointmutationInsertionsordeletionstautomerismonbase-pairingDNApolymoraseIThenatureofmutationsPointmutations:Transitions(pyrimidinetopyrimidine,purinetopurine)Transversions(pyrimidinetopurine,purinetopyrimidine)InsertionsordeletionsATG,GTG,……GAG,CCG,GCC,GAG,TAGATG,GTG,……GAG,CTG,GCC,GAG,TAGATG,GTG,……GAG,CCG,GCC,GAG,TAGATG,GTG,……GAG,CG,GCC,GAG,TAGATG,GTG,……GAG,CGG,CCG,AGT,AGPointmutationInsertionsordeletionsmissenseErrorsinreplicationReplicationerrorsandtheirrepairRepairofReplicationerrors:

Mismatchrepair

IncreasetheaccuracyofDNAsynthesisfor2-3ordersofmagnitudes.Prokaryote

EukaryoteMismatchrepairremoveserrorsthatescapeproofreadingTwochallenges:Rapidlyfindthemismatches/mispairs2AccuratelycorrectthemismatchProkaryoteMismatchrepairsystemMutSMutLMutHUrvDoneofthreeexonucleasesMutSscanstheDNA,recognizingthemismatchfromthedistortiontheycauseintheDNAbackboneMutSembracesthemismatch-containingDNA,inducingapronouncedkinkintheDNAandaconformationalchangeinMutSitselfDNAiskinkedMutSisadimer.Onemonomerinteractswiththemismatchspecifically,andtheothernonspecifically.recruitsMutL,MutLactivatesMutH,enzymecausinganincisionornickononestrandnearthesiteofthemismatch.Nickingisfollowedbythespecifichelicase(UrvD)andoneofthreeexonucleasesDifferentexonucleasesareusedtoremovessDNAbetweenthenickcreatedbyMutHandthemismatch.

recruitsMutL,MutLactivatesMutH,enzymecausinganincisionornickononestrandnearthesiteofthemismatch.Nickingisfollowedbythespecifichelicase(UrvD)andoneofthreeexonucleasesExonucleaseVIIorIDNApolymeraseIIIHowdoestheE.colimismatchrepairsystemknowwhichofthetwomismatchednucleotidetoreplace?DAMmethylasemethylateAresiduesonbothstrandsof5’-GATC-3’ReplicationgeneratehemimethylationDNAinE.coliMutHmakesincisionunmethylateddaughterstrandErrorsinreplicationReplicationerrorsandtheirrepairRepairofReplicationerrors:

Mismatchrepair

IncreasetheaccuracyofDNAsynthesisfor2-3ordersofmagnitudes.Prokaryote

EukaryoteEukaryoticcells

alsorepairmismatchesanddosousinghomologstoMutS(MSH)andMutL(MLH).Theunderlyingmechanismsarenotthesameandnotwellunderstood.

DNAdamageandtheirrepair

CausesRepairDirectreversalofDNAdamageBaseexcisionrepairNucleotideexcisionrepairRecombination(DSB)repairsTranslesionDNAsynthesisdamagespontaneouslalkylation,oxidationandradiationbaseanalogsintercalatingagentsDNAundergoesdamage

spontaneously

fromhydrolysisanddeamination

DeaminationCUHydrolysiscreatesapurinicdeoxyribose

Deamination5-mCT

ExplainingwhyDNAcontainsTinsteadofUAlkylation,OxidationNitrosamines(亞硝胺)Reactiveoxygenspecies(O2-,H2O2,OH?)Radiationbaseanalogs

pointmutation

intercalatingagents

insertionordeletionTwoconsequenceofDNAdamageSomedamages,suchasthyminedimer,nickorbreaksintheDNAbackbone,createimpedimentstoreplicationortranscriptionSomedamagescreatesalteredbasesthathasnoeffectonreplicationbutcausemispairing,whichinturncanbeconvertedtomutation.DNAdamageandtheirrepair

CausesRepairDirectreversalofDNAdamageBaseexcisionrepairNucleotideexcisionrepairRecombination(DSB)repairsTranslesionDNAsynthesisdamagespontaneouslalkylation,oxidationandradiationbaseanalogsintercalatingagentsdamagespontaneouslalkylation,oxidationandradiationbaseanalogsintercalatingagentsDirectreversalofDNAdamagePhotoreactivationMonomerizationofthyminedimersbyDNAphotolyasesinthepresenceofvisiblelight.Methyltransferasecatalyzesthe

transferofthemethylgrouponO6-methylguaninetocysteinresidueontheenzyme,therebyrestoringthenormalGinDNA.MethylgroupremovalBaseExcisionrepairAPendonulease&exonulcease

CleavestheabasicsugarsDNApolymerase/ligase

Workssequentiallytocompletetherepairevent.GlycosylaseRecognizesthedamagedbaseRemovesthedamagedbaseoxoG:Arepair.

AglycosylaserecognizesthemispairandremovesA.

Fail-safesystemsNucleotideExcisionrepairRecognizedistortionstotheshapeoftheDNAdoublehelixRemoveashortsingle-strandedsegmentthatincludesthelesion.DNApolymerase/ligasefillinthegap.NucleotideExcision-RepairUvrA:detectthedistortionofDNAUvrB:meltDNAtocreatessDNArecruitsCUvrCcreatetwoincisionsUvrDreleasethessDNATranscription-couplerepair:nucleotideExcisionrepairsystemiscapableofrescuingRNApolymerasethathasbeenarrestedbythepresenceoflesionsintheDNAtemplateRecombinationrepairsDouble-strandbreak(DSB)re