1、DNA的化學損傷及其修復,1,學習交流PPT,堿 基,2,學習交流PPT,1.DNA的損傷 1.1DNA的自發(fā)損傷：水解和脫氨基作用 1.1.1脫氨基作用 CU U:A AH H:C GX X:C,P269,3,學習交流PPT,1.DNA的損傷 1.1DNA的自發(fā)損傷：水解和脫氨基作用 1.1.2脫堿基,P269,4,學習交流PPT,1.DNA的損傷 1.2DNA的損傷：烷化反應、氧化反應和輻射 1.2.1烷化反應 G甲基化為O6-甲基鳥嘌呤， 與T配對。G:C轉換為A:T 1.2.2氧化反應 鳥嘌呤氧化為氧代鳥嘌呤 (oxoG)，可與A或C配對 G:C顛換為T:A,P269,5,學習交流PP

2、T,1.DNA的損傷 1.2DNA的損傷：烷化反應、氧化反應和輻射 1.2.3輻射 紫外線 電離輻射,P269,6,學習交流PPT,1.DNA的損傷 1.3DNA的損傷：堿基類似物和嵌入劑 堿基類似物 嵌入劑,P271,7,學習交流PPT,2.DNA損傷的直接逆轉 (direct reversal of DNA damage) 2.1光激活作用（photoreactivation） DNA光解酶（DNA photolyase）,P273,8,學習交流PPT,2.DNA損傷的直接逆轉 (direct reversal of DNA damage) 2.2甲基轉移酶（methyltransfera

3、se）,P273,9,學習交流PPT,3.切除修復 (excision repair) 3.1堿基切除修復（base excision repair） 3.2核苷酸切除修復（nucleotide excision repair）,10,學習交流PPT,3.切除修復 (excision repair) 3.1堿基切除修復（base excision repair） DNA糖基化酶識別錯誤堿基并將其除去，形成脫氧戊糖 AP核酸內切酶切除脫堿基戊糖 DNA聚合酶I以未受損的DNA鏈為模板合成新鏈，DNA連接酶連接新鏈,P274,11,學習交流PPT,3.切除修復 (excision repair)

4、3.1堿基切除修復（base excision repair） 3.1.1修復過程 DNA糖基化酶識別錯誤堿基并將其除去，形成脫氧戊糖,P274,12,學習交流PPT,3.切除修復 (excision repair) 3.1.1修復過程（base excision repair） AP核酸內切酶和核酸外切酶切除脫堿基戊糖 DNA聚合酶I以未受損的DNA鏈為模板合成新鏈，DNA連接酶連接新鏈,P274,13,學習交流PPT,3.切除修復 (excision repair) 3.1堿基切除修復（base excision repair） 3.1.1修復過程 DNA糖基化酶識別錯誤堿基并將其除去，形

5、成脫氧戊糖 AP核酸內切酶和核酸外切酶切除脫堿基戊糖 DNA聚合酶I以未受損的DNA鏈為模板合成新鏈，DNA連接酶連接新鏈,P274,14,學習交流PPT,15,學習交流PPT,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 inc

6、ision 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

7、 diesterase, NIR, and 35exonuclease activities,16,學習交流PPT,17,學習交流PPT,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, t

8、hese 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 p

9、athway.,18,學習交流PPT,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.

10、APE1 cleaves oligonucleotide duplexes containing a single UG base pair; this activity depends strongly on the sequence context and the base opposite to U.,19,學習交流PPT,The apparent kinetic parameters of the reactions show that APE1 has high affinity for DNA containing U but cleaves the DNA duplex at a

11、n 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.,20,學習交流PPT,The fact that U in duplex DNA is recognized and cleaved by APE1 in vitro s

12、uggests 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.,21,學習交流PPT,3.切除修復 (excision

13、 repair) 3.1堿基切除修復（base excision repair） 3.1.2損傷堿基彈出,P275,22,學習交流PPT,3.切除修復 (excision repair) 3.1堿基切除修復（base excision repair） 3.1.3防故障機制,What if a damaged base is not removed by base excision before DNA replication?,P275,23,學習交流PPT,3.切除修復 (excision repair) 3.2核苷酸切除修復（nucleotide excision repair） UvrA-UvrB識別并結合損傷堿基所產生的扭曲位點，且UvrA脫離,P275,24,學習交流PPT,3.切除修復 (excision repair) 3.2核苷酸切除修復（nucleotide excision repair） UvrA-UvrB識別并結合損傷堿基所產生的扭曲位點，且UvrA脫離 UvrB使損傷堿基附近DNA變性，產生單鏈凸起 UvrB募集UvrC，UvrC在錯誤堿基兩側切開DNA UvrD出去被切割的DNA片段，Pol和連接酶修補缺口,25,學習交流PPT,3.切除修復 (excision repair) 3.2核苷酸切除修復（nucleotide exci