DNA的化学损伤ppt课件

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 CU U:A AH H:C GX X: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 (direct reversal of DNA damage)2.1光激活作用（photoreactivation）DNA光解酶（DNA photolyase）,P273,2.DNA损伤的直接逆转 (direct reversal of DNA damage)2.2甲基转移酶（methyltransferase）,P273,3.切除修复 (excision repair)3.1碱基切除修复（base excision repair）3.2核苷酸切除修复（nucleotide excision repair）,3.切除修复 (excision repair)3.1碱基切除修复（base excision repair）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 (excision repair)3.1碱基切除修复（base excision repair）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 (excision repair)3.1.1修复过程（base excision repair）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 (excision repair)3.1碱基切除修复（base excision repair）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,The majority of oxidatively damaged DNA bases are substrates for two overlapping pathways: DNA glycosylase-initiated base excision repair (BER) and apurinic/apyrimidinic (AP) endonuclease-mediated nucleotide incision repair (NIR) . In the NIR pathway, an AP endonuclease makes an incision 5 to a damaged nucleotide and then extends the resulting single-strand break to a gap by a nonspecific 35 exonuclease activity. AP endonucleases are multifunctional DNA repair enzymes that possess AP site nicking, 3 repair diesterase, NIR, and 35exonuclease activities,Spontaneous hydrolytic deamination of cytosine to uracil (U) in DNA is a constant source of genome instability in cells. This mutagenic process is greatly enhanced at high temperatures and in single-stranded DNA. If not repaired, these uracil residues give rise to CT transitions, which are the most common spontaneous mutations occurring in living organisms and are frequently found in human tumors. In the majority of species, uracil residues are removed from DNA by specific uracil-DNA glycosylases in the base excision repair pathway.,Alternatively, in certain archaeal organisms, uracil residues are eliminated by apurinic/apyrimidinic (AP) endonucleases in the nucleotide incision repair pathway. Here, we characterized the substrate specificity of the major human AP endonuclease 1, APE1, toward U in duplex DNA. APE1 cleaves oligonucleotide duplexes containing a single UG base pair; this activity depends strongly on the sequence context and the base opposite to U.,The apparent kinetic parameters of the reactions show that APE1 has high affinity for DNA containing U but cleaves the DNA duplex at an extremely low rate. MALDI-TOF MS analysis of the reaction products demonstrated that APE1-catalyzed cleavage of a UG duplex generates the expected DNA fragments containing a 5-terminal deoxyuridine monophosphate.,The fact that U in duplex DNA is recognized and cleaved by APE1 in vitro suggests that this property of the exonuclease III family of AP endonucleases is remarkably conserved from Archaea to humans. We propose that nucleotide incision repair may act as a backup pathway to base excision repair to remove uracils arising from cytosine deamination.,3.切除修复 (excision repair)3.1碱基切除修复（base excision repair）3.1.2损伤碱基弹出,P275,3.切除修复 (excision repair)3.1碱基切除修复（base excision repair）3.1.3防故障机制,What if a damaged base is not removed by base excision before DNA replication?,P275,3.切除修复 (excision repair)3.2核苷酸切除修复（nucleotide excision repair）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 (excision repair)3.2核苷酸切除修复（nucleotide excision repair）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 (excision repair)3.