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ID:40230969
大小:15.20 MB
页数:62页
时间:2019-07-27
《04-2 Biosynthesis of DNA--Replication》由会员上传分享,免费在线阅读,更多相关内容在行业资料-天天文库。
1、§4TheBiosynthesisofDNA---Replication(2)DNAsynthesisrequiresdeoxynucleosidetriphosphatesandaprimer:templatejunctionDNAissynthesizedbyextendingthe3’endoftheprimerHydrolysisofpyrophosphate(PPi)isthedrivingforceforDNAsynthesisReviewSubstraterequiredforDNAsynthesis§4-5Enzymology
2、ofDNAReplicationReplisomeisthemultiproteinstructurethatassemblesatthereplicationforktoundertakesynthesisofDNA.ContainsDNApolymeraseandotherenzymes.1.DoubleStrandsSeparationItisthefirststepofDNAreplicationthatdoublestrandsseparateintosinglestrand.Threekindsofproteinareimport
3、antinthisprocess.1)Helicase:AhelicaseisanenzymethatseparatesthestrandsofDNA,usuallyusingthehydrolysisofATPtoprovidethenecessaryenergy.E.colihelicase:DnaB,PriAandRepprotein.Helicasealsojoinsintheprocessofrepairing.2)Single-strandbindingprotein(SSB):Asingle-strandbindingprote
4、in(SSB)bindstothesingle-strandedDNA,preventingitfromreformingtheduplexstate.TheSSBbindsasamonomer,buttypicallyinacooperativemannerinwhichthebindingofadditionalmonomerstotheexistingcomplexisenhanced.SSBalsojoinsintheprocessofrepairing.3)Topoisomerase(gyrase):负责产生负超螺旋并使两个子代环状
5、DNA分子分开。Single-strandedbindingproteins(SSBs)stabilizesingle-strandedDNA2.DNApolymeraseAnenzymethatcansynthesizeanewDNAstrandonatemplatestrandiscalledaDNApolymerase.ProkaryticDNApolymeraseE.coliDNApolymeraseⅠ,proofreadingremovesRNAprimersⅡ,repairingⅢ,replicationIV,SOSV,SOS1)
6、E.coliDNApolymeraseE.coliDNApolymeraseI:monomer,109kDa,928AA。用枯草杆菌酶或胰蛋白酶消化,可获得大小两个片段:Largefragment--KlenowI:C端324~928AA,5’-3’polymeraseand3’-5’exonuclease.Smalldomain:5’-3’exonucleaseProofreadingNicktranslationProofreading:Proofreadingreferstoanymechanismforcorrectingerrors
7、inproteinornucleicacidsynthesisthatinvolvesscrutinyofindividualunitsaftertheyhavebeenaddedtothechain.3’-5’exonuclease5’-3’polymeraseExonucleasesproofreadnewlysynthesizedDNATheoccasionalflickingofthebasesinto“wrong”tautomericformresultsinincorrectbasepairandmis-incorporatio
8、nofdNTP.(10-5mistake)ThemismatcheddNMPisremovedbyproofreadingexonuclease,apartofth
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