DNA損傷和修復(fù)

1、Chapter5DNAdamageandrepair1 .主要內(nèi)容1）誘變2 ）DNA損傷3 ）DNA修復(fù)4 .教學(xué)要求：1）熟悉DNA損傷的原因、類型2）掌握DNA修復(fù)的方式5.1 Mutagenesis5.2 DNAdamage5.3 DNARepair5.1 Mutagenesis例變）?5.1.1Thereasonsofmutation?5.1.2Typesofmutations?5.1.3Mutagens傍變劑）?5.1.4mutagenesis誘變）5.1 Mutagenesis（誘變）?Mutation（突變）=Permanent,heritablealterationsinth

2、ebasesequenceofaDNAmolecule（是DNA堿基序列水平上永久性的、可遺傳的變化）5.1.1 Thereasonsofmutation?Spontaneouserrors自發(fā)性錯(cuò)誤）:inDNAreplicationormeioticrecombination?Mutagen（誘變物）:AconsequenceofthedamagingeffectsofphysicalorchemicalmutagensonDNA?Mutagen=mutationcausingagent?Essentiallyallmutagensarecarcinogen斂癌物）?Mostcarcino

3、gensaremutagens5.1.2 Typesofmutations?Multisite（多位點(diǎn)）：- Causegross（重的）chromosomeabnormalities（奇形）；-InvolvelargeregionsofDNA；-Ariseduringmeiosis（在減數(shù)分裂期發(fā)生）?Pointmutations（點(diǎn)突變）-Involveonlyoneorafewnucleotides；- AriseduringDNAreplication- Requiretwoerrors:1）AnerrorduringDNAreplication；2）Failuretocorrectt

4、haterrorTypesofpointmutationsInversions:ACBDEFDuplicationsABCDEEFDeletions:ABCD-FInsertions:ABCDSEFSubstitutions:ATCDEFTypesofpointmutationsThephenotypiceffectofpointmutation（點(diǎn)突變的表型效應(yīng)）?Missensemutation（昔義突變）：堿基序列的改變引起了產(chǎn)物氨基酸序列的改變。?Nonsensemutation（無(wú)義突變）：某個(gè)堿基的改變使代表某種氨基酸的密碼子變?yōu)榈鞍踪|(zhì)合成的終止密碼子（stopcodon,TAG,

5、TAA,TGA）。?Samesensemutation（同義突變）：是指未改變產(chǎn)物氨基酸序列的密碼子變化。Noeffect（silentmutation沉默突變）Readingframe（閱讀框）isoneofthethreepossiblewaysinwhichanmRNAsequenceofnucleotidescanbereadasaseriesofbasetriplets三個(gè)一組tospecifytheaminoacidsinaproteinchain.ORF（openreadingfram/放性I閱讀框）:fromstartcodontostopcodonWhatisthefirst

6、defenseagainstmutations?Ontheonehand,theactualerrorrateofthepolymerase,beforeediting,isoftheorderof0.1%to1.0%.However,theoverallerrorrateforDNAreplicationis1errorin109to1010basepairs.Thisphenomenalfidelityisachievedinthreeways.Replicationfidelity（復(fù)制的保真度）First,Watson-CrickbasepairingtoSecond,DNApolym

7、eraseshavetheabilitytoedit（"proofread"）theirwork（3exonucleaseactivityofthepolymerases）.Third,post-replicationrepairofDNA（mismatchrepair）.5.1.3 Mutagens例變劑）Physicalmutagens（物理誘變齊J）?High-energyionizingradiation（電離輻射）：X-raysand-raysstrandbreaks斷鏈），base/sugardestruction（堿基/核糖損傷）?Nonionizingrad

8、iation（非電離輻射）：UVlightpyrimidinedimers（示呢二聚體）Chemicalmutagens（化學(xué)誘變劑）?Baseanalogs（堿基類彳以物）:mispair,directmutagenesis?Nitrousacid（亞硝酸）：deaminatesCtoproduceU（脫氨基作用）?Alkylatingagents（烷化劑）?Arylatingagents（芳基化劑）5.1.4 Mutagenesis例變）?Directmutagenesis直接誘變）：resultsfromthepresenceofstable,unrepairedbasewithalte

9、redbasepairingpropertiesintheDNA.Indirectmutagenesis（同接誘變）:Themutationisintroducedasaresultofanerror-pronerepair（傾向差錯(cuò)的修復(fù)）.一Translesion的越損傷）DNAsynthesis:Insertionofbasesoppositetheunrepairedlesionregardlessoftheoriginalsequence（不顧原始序列如何，在未修復(fù)的損傷序列對(duì)面插入堿基）-tomaintaintheDNAintegritybutnotthesequenceaccur

10、acy（只彳於證DNA序列的完整性，不保證序列的精確性）5.2 DNAdamageDNAlesions損傷：oxidativedamages化性損傷Alkylation烷基化bulkyadducts聚化力口合物5.2.1 DNAlesions（DNA損傷）?DNAlesions（DNA損傷）：AnalterationtothenormalchemicalorphysicalstructureoftheDNA（DNA正常的化學(xué)或物理結(jié)構(gòu)的改變）5.2.3 Alkylation（烷基化作用）5.2.4 Bulkyadducts（聚化加合物）Bulkylesionssuchaspyrimidined

11、imersandarylating芳基化agentadductsdistortthedoublehelixandcauselocalized局部denaturation.ThisdisruptsthenormalfunctioningoftheDNA.口密噬二聚體的形成喀呢二聚體是如何造成DNA損傷的？?由于相鄰的胸腺喀呢產(chǎn)生二聚體，兩個(gè)堿基平面被環(huán)丁基所扭轉(zhuǎn)，引起雙螺旋構(gòu)型的局部變化，同時(shí)氫鍵結(jié)合力也顯著減弱。這樣，當(dāng)有胸腺喀呢二聚體的DNA作為模板進(jìn)行復(fù)制時(shí)，Polin將兩個(gè)腺喋吟核甘酸加上去，但由于不能很好地形成氫鍵，然后，又由3'-5'校對(duì)功能而將之水解。如此反復(fù)發(fā)生，

12、而產(chǎn)生了一個(gè)空耗的過程，即大量的dATP被分解，而DNA復(fù)制卻毫無(wú)進(jìn)展。喀呢二聚體是如何造成DNA損傷的？?由于蛋白質(zhì)仍在不斷合成，而DNA不能復(fù)制，細(xì)胞也就不能分裂，結(jié)果出現(xiàn)細(xì)絲狀的所謂蛇形細(xì)胞，最后導(dǎo)致細(xì)胞死亡。5.3 DNARepair（DNA修復(fù)）?DirectRepair：-Photoreactivation（光復(fù)活）-Alkyltransferase（烷基轉(zhuǎn)移酶）?Exisionrepair（切除修復(fù)）?Mismatchrepair（錯(cuò)配彳復(fù)）?Post-ReplicationRepair（復(fù)制后修復(fù)）-recombinationalrepair（重組修復(fù)）-SOSrepair5.

13、3.1 Photoreactivation洸復(fù)活）在可見光的存在下，DNA光解酶（photolyase,光復(fù)活酶）可將環(huán)丁烷二聚體再分解為單體。這些酶含有可吸收藍(lán)光并將能量轉(zhuǎn)移到待切環(huán)丁烷環(huán)中的輔基。E.coli的光解酶含有2個(gè)色素分子，N5,N10-次甲基四氫葉酸和還原性的黃素腺喋吟二核甘酸（FAD）0光復(fù)活對(duì)喀呢二聚體是專一性的。是損傷被直接修復(fù)”的一種例子，是無(wú)差錯(cuò)的。PhotoreactivationofThymineDimers?Processiscatalyzed®化inaprocesssimilartophotosynthesis-harvestingenergyfro

14、mlight?Humancellsdonotcontainphotolyase5.3.2 Alkyltransferase皖基轉(zhuǎn)移酶）?AlkylationofDNABases:?Canblock®礙DNAreplicationbecauseofmodifiedbasesthatareformed?Sometimeusedinchemotherapy（化學(xué)療法）toblockcelldivision?Usuallypurinesarealtered-spectrurtt譜ofproductsvariesMosthighlymutagenicoftheseproducts:Guanin

15、eO6-alkylguanine錯(cuò)配的O6-methylguanin琲口thymine一起造成GC替換為AT。無(wú)差錯(cuò)直接修復(fù)?烷基轉(zhuǎn)移酶特異性地轉(zhuǎn)移O6耳基鳥口S吟或O6-乙基鳥喋吟上的甲基或乙基基團(tuán)到酶分子的半胱氨酸上，從而保護(hù)DNA免受烷基化誘變。5.3.3 Exisionrepair（切除彳復(fù)）?Including一NucleotideExcisionRepair（NER,核甘酸切除修復(fù)）E.coliUvrABCendonuclease系統(tǒng)一Baseexcisionrepair（BER堿基切除修復(fù)）-Uracil-DNANglycosylasesystem（糖基化酶系統(tǒng)）?Isaubiq

16、uitousmechanismrepairingavarietyoflesions是修復(fù)多種損傷的普遍性機(jī)制?Error-freerepai吮差錯(cuò)修復(fù)核甘酸切除修復(fù)ExcisionrepairsystemsinE.colil.uvrA和uvrB蛋白識(shí)別損傷部位。2 .發(fā)現(xiàn)二聚體后，uvrA解離，uvrC加入。3 .uvrB和uvrC復(fù)合體在損傷部位的兩端造成單鏈缺口。4 .損傷部位的DNA被DNA螺旋酶（uvrD）移走。5 .DNA上的缺口由DNA聚合酶I和連接酶填補(bǔ)）。核甘酸切除修復(fù)ExcisionrepairsystemsineukaryotesNucleotideexcisionrepa

17、irxerodermapigmentosum（XP,著色性干皮?。┦且环N常染色體隱性紊亂，在表型上表現(xiàn)為對(duì)陽(yáng)光極為敏感，極易產(chǎn)生皮膚癌。XP患者缺乏對(duì)包括由紫外線引起的大塊DNA損傷的核甘酸切除功能，至少有7種不同基因的缺陷可導(dǎo)致XP。Baseexcisionrepair（BER,堿基切除修復(fù)）5.3.4 Mismatchrepair仔音配彳修復(fù)）?用于修復(fù)在復(fù)制中錯(cuò)配并漏過校正檢驗(yàn)的任何堿基。可使復(fù)制的保真性提高102-103倍。?occursjustafterreplication?mustdistinguishtheparentfromthedaughterstrand5.3.5 Rec

18、ombinationalRepair（重組修復(fù)）?Presentinprokaryoticandeukaryoticcells?Onlypoorlyunderstood?WeknowitexistsbecauseUvrA-andRecA-cellsaremuchmoresensitivetoUVthancellscontainingonlyonemutation.RecombinationalRepair?子鏈在母鏈損傷處形成缺口?DependsonRecAproteinimportantforrecombinationandrepair;itcatalyzesstrandpairing.?以

19、子鏈為模板填補(bǔ)母鏈的缺口?復(fù)制重新開始，損傷位點(diǎn)被切除修復(fù)5.3.6 SOSrepair（SOS修復(fù)）?UVreactivation（紫外激活反應(yīng)）或Wreactivation（W激活反應(yīng)）：-50年代J.Weigle發(fā)現(xiàn)，用經(jīng)紫外線照射后的噬菌體感染用低劑量UV照射過的大腸桿菌時(shí)，噬菌體的存活率比感染未用低劑量UV照射的大腸桿菌明顯增加，突變率也隨之增加，這一效應(yīng)稱為UVreactivation（紫外激活反應(yīng)）或Wreactivation（W激活反應(yīng)）。?TheincreasedsurvivalintheUVirradiatedhostisduetotheinductionoftheSOS-

20、repairsysteminthehost.（存活率上升是由于紫外線照射引起了寄主SOS修復(fù)系統(tǒng)的感應(yīng)）?SOSt復(fù)是一種旁路系統(tǒng),它允許新生的DNA鏈越過胸腺喀呢二聚體而生長(zhǎng)，其代價(jià)是保真度的極大降低，這是一種易錯(cuò)修復(fù)系統(tǒng)。SOS反應(yīng)機(jī)制?Metabolicsystemthathelpsthecellsurviveinperiodsofpotentiallylethalstresse的潛在的致死壓力下，細(xì)胞的新陳代謝系統(tǒng)幫助細(xì)胞存活）-Inducedby（誘導(dǎo)因素）：?UVirradiation（紫外照射）?Thyminestarvation（胸腺喀呢饑餓）?TreatmentwithDNAmodifyingenzymes（DNA修飾酶處理）?Inactivationofgenesessentialtoreplication（DNA復(fù)制必須的基因失活）TheSOSgenesSOSrepairDiseasescausedbydefectsinRecQhelicasegenesWernersyndrome?prematureagin（gi衰老,beginningin20's;lifeexp.45?Predisposition患病白體質(zhì)tomalignanciesDiseasescausedbydefectsinRecQhelicasegen