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SECTIONFDNAdamage,repair,andrecombination•MolecularBiologyCourseF1Mutagenesis(诱变)Mutation:replicationfidelity,mutagens,mutagenesisF2DNAdamageDNAlesions:oxidativedamage,alkylation,bulkyadductsF3DNArepairPhotoreaction,alkyltransferase,excisionrepair,mismatchrepairF4RecombinationHomologousrecombination,site-specificrecombination,transpositionF1Mutagenesis(诱变)1.Mutation(突变)2.Replicationfidelity3.Mutagens(诱变剂):chemical&physical4.Mutagenesis:direct&indirectDNAdamage,repair,andrecombinationDNAdamage,repair,andrecombinationPermanent,heritablealterationsinthebasesequenceofDNAReasons1.SpontaneouserrorsinDNAreplicationormeioticrecombination2.AconsequenceofthedamagingeffectsofphysicalorchemicalmutagensonDNAF1-1Mutation(突变)Transition(转换):嘌呤与嘌呤、嘧啶与嘧啶A—GT—CTransversion(颠换):嘌呤与嘧啶互换A—TorCT—AorGG—TorCC—AorGPointmutation(点突变)(asinglebasechange)•NoncodingDNA•NonregulatoryDNA•3rdpositionofacodonEffectsofapointmutationSilentmutation(沉默突变)PhenotypiceffectsNoMissensemutation(错义突变)CodingDNAalteredAAYesorNoCodingDNAstopcodonruncatedproteinNonsensemutation(无义突变)YesInsertionsordeletions一个或多个碱基的增加或缺失Frameshiftmutations(移码突变)结果是所翻译出的蛋白质序列从突变点起至C末端都完全被改变DNAdamage,repair,andrecombinationF1-2ReplicationfidelityImportantforpreservethegeneticinformationfromonegenerationtothenextMutationrelevant1.SpontaneouserrorsinDNAreplicationisveryrare,oneerrorper1010baseinE.coli.F1Mutagenesis(诱变)1.DNApolymerase:Watson-Crickbasepairing2.3’-5’proofreadingexonuclease.3.RNApriming:proofreadingthe5’endofthelaggingstrand4.Mismatchrepair(错配修复)(F3)MolecularmechanismsforthereplicationfidelityF1-3&4:Mutagens(诱变剂)MutationrelevantCauseDNAdamagethatcanbeconvertedtomutations.DNAdamage,repair,andrecombinationPhysicalmutagens(物理诱变剂)High-energyionizingradiation:X-raysNonionizingradiation:UVlightF1-3&4:Mutagens(诱变剂)其实X射线在我们的日常生活中是无处不在的。电视、电脑、手机、微波炉,就连大自然赐予人类的太阳光和月亮光线也存在着X射线。而日常生活中有些物体的X射线危害也是不能忽视的,例如不要让小孩子太接近家中正在工作中的微波炉;选用建筑材料时要仔细挑选,像辐射较大的花岗石就要慎用;平时也不要长时间看电视或使用电脑,如果是工作所需,尽可能做到工作一、两个小时就让自己休息十分钟。另外,尽量不要连续在短期内做多次X射线检查,如果做了,就要注意吃些维生素补充一下,例如猕猴桃,海带,红薯等.Chemicalmutagens(化学诱变剂)Baseanalogs(碱基类似物)Nitrousacid(亚硝酸)Alkylatingagents(烷化剂)F1-3&4:Mutagens(诱变剂)F1-3&4:Mutagenesis(诱变)Themolecularprocessinwhichthemutationisgenerated.Directmutagenesis(直接诱变)Indirectmutagenesis(间接诱变)DNAdamage,repair,andrecombination起因于DNA中存在稳定的、具改变了的配对特性的未修复的碱基。DirectmutagenesisDNAdamage,repair,andrecombinationDNA的复制使得这种损伤固定下来成为永久的,可遗传的。5’溴尿嘧啶-T类似物烯醇式酮基异构体间接诱变损伤发生在移动的复制叉之前,不可能修复.|子链产生致命的缺口|转移损伤DNA合成-随便插入碱基,容忍错误间接诱变:损伤DNA聚合酶只是为了保证染色体的完整性在损伤对应的位点插入错误的碱基。F1Mutagenesis(诱变)1.Mutation(突变)2.Replicationfidelity3.Mutagens(诱变剂):chemical&physical4.Mutagenesis:direct&indirectDNAdamage,repair,andrecombination诱变剂自发-碱基化学失活小的温和的改变完全修复错配修复复制叉之前突变间接诱变直接诱变F2DNA损伤致死F3修复F2DNAdamageDNA损伤氧化性损伤烷基化聚化加合物•DNAdamage,repair&recombinationF2-1DNAlessionsAnalterationtothenormalchemicalorphysicalstructureoftheDNAF2DNAdamage外源物质:化学试剂射线致突变和致死自发性1.Deamination(转氨作用):C-U2.Depurination(脱瞟呤作用):3.Depyrimidine(脱嘧啶作用)F2DNAdamageF2-2Oxidativedamage氧化性损伤定义:在所有需氧细胞中由于超氧化物、氢过氧化物及最重要的羟基自由基等活性氧的存在,会在正常条件下攻击DNA,产生一系列特性变化了的氧化产物。F2DNAdamageF2-3Alkylation烷基化烷化剂可以将烷基加入到核酸上的各种位点。这些损伤中的部分会在DNA复制及转录时干扰DNA解旋F2DNAdamageF2-4Bulkyadducts聚化加合物嘧啶二聚体TheselesionsdisruptthenormalfunctionoftheDNA-不能正常配对F2DNAdamageDNAlesionsthatdistortthedoublehelixandcauselocalizeddenaturation,煤焦油中致癌物;黄曲霉毒素B1F3DNArepairPhotoreactivation(光复活作用)Alkyltransferase(烷基转移酶)Exisionrepair(切割修复)Mismatchrepair(错配修复)DNAdamage,repair,andrecombinationPhotoreactivation(光复活)在可见光存在的情况下,DNA光解酶(光复活酶)可将环丁烷嘧啶二聚体分解为单体。Directreversalofalesionandiserror-freeF3DNArepairAlkyltransferase(烷基转移酶)Thealkylgroupistransferredtotheproteinitselfandinactivateit.F3DNArepair烷基转移酶能直接从突变的O6-烷基鸟嘌呤上去除烷基。-致死性保护Exisionrepair(切除修复)核苷酸切除修复(NER)碱基切除修复(BER)F3DNArepair核苷酸切除修复(NER)内切核酸酶在损伤部位两边各切除精确数目的碱基UvrABC内切核酸酶-嘧啶二聚体损伤部位切除,留下缺口DNA聚合酶DNA连接酶大损伤F3DNArepair-Exisionrepair(切除修复)碱基切除修复(BER)DNA糖基化酶识别修饰碱基,切除糖苷键,留下脱嘌呤或脱嘧啶(AP)/自发碱基丢失AP内切核酸酶在该位点切开,其外切核酸酶活性继续切除一个缺口DNA聚合酶,DNA连接酶B*AP修饰碱基F3DNArepairF3DNArepair-Exisionrepair(切除修复)Mismatchrepair(错配修复)错配修复是切除修复的一种特定形式,用于修复在复制中错配并漏过校正检验的任何碱基。F3DNArepairMutS和MutL蛋白复合体识别错配碱基MutH内切核酸酶与之结合,在子代链特异性的产生缺刻DNA聚合酶,DNA连接酶半甲基化:亲本链甲基化,子代链未被甲基化(正常GACT中A甲基化)F3DNArepair-Mismatchrepair(错配修复)错配修复机制F4Recombination(重组)1.Homologousrecombination2.Site-specificrecombination3.Transposition(转座)DNAdamage,repair,andrecombination1.Homologousrecombination同源重组也称一般重组,涉及两个DNA分子间同源区域的交换。双倍体真核生物:减数分裂过程单倍体原核生物:Holliday模型DNAdamage,repair,andrecombination双倍体真核生物-同源重组1.Homologouschromosomeslineupinmeiosis(when)2.Thenonsisterchromatidsexchangeequivalentsections(what)DNAdamage,repair,andrecombination单倍体原核生物-同源重组(E.coli)(where)1.发生在部分已复制DNA之间2.发生在染色体DNA与获取的外源DNA之间BetweenthetwohomologousDNAduplexHollidaymodel(How)DNAdamage,repair,andrecombinationrecA-dependent细菌同源重组同源DNA序列1.核酸酶和RecBCD蛋白复合体在特定序列chi产生缺口2.含有5’端切口的单链DNA被RecA蛋白包裹形成RecA-ssDNA细丝3’3’5’5’3.细丝交换,寻找相对的DNA双螺旋上的相应区域(侵入)4.切口被封好,形成四分支的Holliday结构5.分支迁移DNAdamage,repair,andrecombination6.解决Holliday拆分fggABbaABabfABabAbaB复制后修复机制a.ReplicationencounteraDNAlesionb.Skipthelesion&reinitiateonthesid
本文标题:分子生物学SectionF
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