DNA的化学损伤ppt课件.ppt

DNA的化学损伤及其修复,.,碱基,1.DNA的损伤1.1DNA的自发损伤：水解和脱氨基作用1.1.1脱氨基作用CUU:AAHH:CGXX:C,P269,1.DNA的损伤1.1DNA的自发损伤：水解和脱氨基作用1.1.2脱碱基,P269,1.DNA的损伤1.2DNA的损伤：烷化反应、氧化反应和辐射1.2.1烷化反应G甲基化为O6-甲基鸟嘌呤，与T配对。G:C转换为A:T1.2.2氧化反应鸟嘌呤氧化为氧代鸟嘌呤(oxoG)，可与A或C配对G:C颠换为T:A,P269,1.DNA的损伤1.2DNA的损伤：烷化反应、氧化反应和辐射1.2.3辐射紫外线电离辐射,P269,1.DNA的损伤1.3DNA的损伤：碱基类似物和嵌入剂碱基类似物嵌入剂,P271,2.DNA损伤的直接逆转(directreversalofDNAdamage)2.1光激活作用（photoreactivation）DNA光解酶（DNAphotolyase）,P273,2.DNA损伤的直接逆转(directreversalofDNAdamage)2.2甲基转移酶（methyltransferase）,P273,3.切除修复(excisionrepair)3.1碱基切除修复（baseexcisionrepair）3.2核苷酸切除修复（nucleotideexcisionrepair）,3.切除修复(excisionrepair)3.1碱基切除修复（baseexcisionrepair）DNA糖基化酶识别错误碱基并将其除去，形成脱氧戊糖AP核酸内切酶切除脱碱基戊糖DNA聚合酶I以未受损的DNA链为模板合成新链，DNA连接酶连接新链,P274,3.切除修复(excisionrepair)3.1碱基切除修复（baseexcisionrepair）3.1.1修复过程DNA糖基化酶识别错误碱基并将其除去，形成脱氧戊糖,P274,3.切除修复(excisionrepair)3.1.1修复过程（baseexcisionrepair）AP核酸内切酶和核酸外切酶切除脱碱基戊糖DNA聚合酶I以未受损的DNA链为模板合成新链，DNA连接酶连接新链,P274,3.切除修复(excisionrepair)3.1碱基切除修复（baseexcisionrepair）3.1.1修复过程DNA糖基化酶识别错误碱基并将其除去，形成脱氧戊糖AP核酸内切酶和核酸外切酶切除脱碱基戊糖DNA聚合酶I以未受损的DNA链为模板合成新链，DNA连接酶连接新链,P274,ThemajorityofoxidativelydamagedDNAbasesaresubstratesfortwooverlappingpathways:DNAglycosylase-initiatedbaseexcisionrepair(BER)andapurinic/apyrimidinic(AP)endonuclease-mediatednucleotideincisionrepair(NIR).IntheNIRpathway,anAPendonucleasemakesanincision5toadamagednucleotideandthenextendstheresultingsingle-strandbreaktoagapbyanonspecific35exonucleaseactivity.APendonucleasesaremultifunctionalDNArepairenzymesthatpossessAPsitenicking,3repairdiesterase,NIR,and35exonucleaseactivities,Spontaneoushydrolyticdeaminationofcytosinetouracil(U)inDNAisaconstantsourceofgenomeinstabilityincells.Thismutagenicprocessisgreatlyenhancedathightemperaturesandinsingle-strandedDNA.Ifnotrepaired,theseuracilresiduesgiverisetoCTtransitions,whicharethemostcommonspontaneousmutationsoccurringinlivingorganismsandarefrequentlyfoundinhumantumors.Inthemajorityofspecies,uracilresiduesareremovedfromDNAbyspecificuracil-DNAglycosylasesinthebaseexcisionrepairpathway.,Alternatively,incertainarchaealorganisms,uracilresiduesareeliminatedbyapurinic/apyrimidinic(AP)endonucleasesinthenucleotideincisionrepairpathway.Here,wecharacterizedthesubstratespecificityofthemajorhumanAPendonuclease1,APE1,towardUinduplexDNA.APE1cleavesoligonucleotideduplexescontainingasingleUGbasepair;thisactivitydependsstronglyonthesequencecontextandthebaseoppositetoU.,TheapparentkineticparametersofthereactionsshowthatAPE1hashighaffinityforDNAcontainingUbutcleavestheDNAduplexatanextremelylowrate.MALDI-TOFMSanalysisofthereactionproductsdemonstratedthatAPE1-catalyzedcleavageofaUGduplexgeneratestheexpectedDNAfragmentscontaininga5-terminaldeoxyuridinemonophosphate.,ThefactthatUinduplexDNAisrecognizedandcleavedbyAPE1invitrosuggeststhatthispropertyoftheexonucleaseIIIfamilyofAPendonucleasesisremarkablyconservedfromArchaeatohumans.Weproposethatnucleotideincisionrepairmayactasabackuppathwaytobaseexcisionrepairtoremoveuracilsarisingfromcytosinedeamination.,3.切除修复(excisionrepair)3.1碱基切除修复（baseexcisionrepair）3.1.2损伤碱基弹出,P275,3.切除修复(excisionrepair)3.1碱基切除修复（baseexcisionrepair）3.1.3防故障机制,WhatifadamagedbaseisnotremovedbybaseexcisionbeforeDNAreplication?,P275,3.切除修复(excisionrepair)3.2核苷酸切除修复（nucleotideexcisionrepair）UvrA-UvrB识别并结合损伤碱基所产生的扭曲位点，且UvrA脱离,P275,3.切除修复(excisionrepair)3.2核苷酸切除修复（nucleotideexcisionrepair）UvrA-UvrB识别并结合损伤碱基所产生的扭曲位点，且UvrA脱离UvrB使损伤碱基附近DNA变性，产生单链凸起UvrB募集UvrC，UvrC在错误碱基两侧切开DNAUvrD出去被切割的DNA片段，Pol和连接酶修补缺口,3.切除修复(excisionrepair)3.2核苷酸切除修复（nucleoti