2009年分子生物学quiz1参考答案(精)

2009年分子生物学quizl参考答案Whatiscentraldogma?Whatcontentshaveyoulearnedineachpartofthecentraldogma?(5’)Thecentraldogmaisthepathwayfortheflowofgeneticinformation.间是双键，CGH一间是双键，CGH一r键；AG是二环，TC是单环；氢键的位ThefeaturesofDNAstructure：Wehavelearntthemaintenanceofthegenome(thestructureofthegeneticmaterialanditsfaithfulduplication)andtheexpressionofthegenome(theconversionofgeneticinstructionscontainedinDNAintoproteins).(缺reversetranscription扣一分)Writeoutthestructureoffourbases,labelthepositionsthatparticipateWatson-Crickbasepairingandthatconnecttotheribose?(10’)如果解答中可以置等，会酌情wg/ qsugarWhatarethefeaturesofDNAstructure?AndhowRNAstructurediffersfromDNA?(2Copyright©2004PearsonEducationfInc.,publishingasBenjaminCummingsTwoantiparallelpolynucleotidechainsaretwistingaroundeachotherintheformofadoublehelix.6’(polynucleotide(buildingblock)2’,antiparallel2’,doublehelix2’)HydrogenBondingdeterminestheSpecificityofbasepairing(complimentarity).2’StackinginteractionbetweenbasesdeterminesthestabilityofDNAdoublehelix,hydrogenbondalsocontributestostability.2’ThedoublehelixhasMinorandMajorgrooves.(ABZforms)2’Differences：Primarystructure(buildingblock):2’dNTPvs.rNTP;Tvs.U;double-strandedvs.single-stranded.Secondarystructure:4’DNAhasstabledoublehelicalstructure.(fullcomplimentarity)RNAchainsfoldbackonthemselvestoformlocalregionsofdoublehelixsimilartoA-formDNA.RNAhelixarethebase-pairedsegmentsbetweenshortstretchesofcomplementarysequences,whichadoptoneofthevariousstem-loopstructures,pseudoknots.(inter-andintra-molecularbasepairing)Tertiarystructures:2’DNA:notertiarystructure.RNAcanfoldupintocomplextertiarystructures,becauseRNAhasenormousrotationalfreedominthebackboneofitsnon-base-pairedregions.ComparethechemistryofDNAsynthesisandRNAsynthesis.(5’)(clues:compare=differences+thesame;chemistry=substrate+direction+energy)Differences:(1)DNAsynthesisrequiresdeoxynucleotidetriphosphateswhileRNAsynthesisrequiresoxynucleotidetriphosphates;(1')(2)ThebaseforDNAsynthesisareA/T/C/GwhilethatforRNAsynthesisareA/U/C/G;(1')(3)DNAsynthesisneedsaprimer:templatejunctionwhileRNAsynthesisdonot;(1')Thesame:(1)ThedirectionofbothDNAsynthesisandRNAsynthesisis5'to3';(1')(2)TheenergyneededforDNAsynthesisaswellasRNAsynthesisishydrolysisofpyrophosphate(PPi).(1')DescribethefunctionsofeachdomainoftheDNApolymerase.(14’+1')DNApolymerasepalmdomain(1'):(1)Containstwocatalyticsites,oneforadditionofdNTPs(1')andoneforremovalofthemispaireddNTP.(1')(2)Thepolymerizationsite:(1)bindstotwometalionsthatalterthechemicalenvironmentaroundthecatalyticsiteandleadtothecatalysis.(1')(2)Monitorstheaccuracyofbase-pairingforthemostrecentlyaddednucleotidesbyformingextensivehydrogenbondcontactswithminorgrooveofthenewlysynthesizedDNA.(1')(3)Exonucleasesite/proofreadingsite.Themechanismofproofreadingiskineticselectivity(1')andthemismatcheddNMPisremovedbyproofreadingexonucleaseinthedirectionof3'-5'(1').DNApolymerasefingerdomain:(1')(1)BindstotheincomingdNTP,enclosesthecorrectpaireddNTPtothepositionforcatalysis;(1')(2)Bendsthetemplatetoexposetheonlynucleotideatthetemplatethatreadyforformingbasepairwiththeincomingnucleotide;(1')(3)Stabilizationofthepyrophosphate;(1')DNApolymerasethumbdomain:(1')(1)Notdirectlyinvolvedincatalysis;(2)InteractswiththesynthesizedDNAtomaintaincorrectpositionoftheprimerandtheactivesite,(1')andtomaintainastrongassociationbetweenDNAPolanditssubstrate.(1')HowreplicationofaDNAmoleculeisaccomplishedinbacteria?

Initiation:RecognitionandbindingofOriCbyDnaA(initiator)-ATP.Helicase(DnaB)loadingandDnaC(helicaseloader)andDNAunwinding.Elongation:PrimasesynthesizesRNAprimer,andDNAPolymeraseIIIsynthesizesthenewDNAstrand.Elongation:(The"trombone"modelwasdevelopedtoexplainlaggingstrandandleadingstrandsynthesizedsimultaneously):Leadingstrand:newlysynthesizedDNAstrandthatiscontinuouslycopiedfromthetemplatestrandbyaDNApolymeraseafterthefirstRNAprimerwasmadebyaprimase.AslidingclampisusuallyloadedtotheDNApolymerasetoincreasethepolymeraseprocessivity.The3directionoftheingstrandisthesameasthemovingdirectionofthereplicationfork.Laggingstrandisdiscontinuouslycopiedfromthetemplatestrand.The3directionofthelaggingstrandisoppositetothemovingdirectionofthereplicationfork.PrimasemakesRNAprimersperiodicallyafterthetemplatestrandisunwoundandbecomessingle-stranded.DNApolymeraseextendseachprimertosynthesisshortDNAfragments,calledOkazakifragments.ThepolymerasedissociatesfromthetemplatestrandwhenitmeetsthepreviousOkazakifragment.Finally,RNAprimersaredigestedbyanRNaseHactivity,andthegapsarefilledbyDNApolymerase.Atlast,theadjacentOkazakifragmentsarecovalentlyjoinedtogetherbyaDNAligasetogenerateacontinuous,intactstrandofnewDNA.Termination:TypeIItopoisomerasesseparatedaughterDNAmolecules.8.HowtranscriptionofaRNAmoleculebyRNApolymeraseIIisinitiated,elongatedandterminatedineukaryotes(25’)Initiation:(11’+附加分)1.Promoterrecognition:TBPinTFIIDbindstotheTATAbox；(1’)TFIIAandTFIIBarerecruitedwithTFIIBbindingtotheBRE；(1’).RNAPolIIrecruitment:RNAPolII-TFIIFcomplexistherecruited；(1’).TFIIEandTFIIHthenbindupstreamofPolII(toformthepre-initiationcomplex).(1()如果提到了pre-initiationcomplex,有1分的附加分)PromotermeltingusingenergyfromATPhydrolysisbyTFIIH.(2’)PromoterescapesafterthephosphorylationoftheCTDtail.(2’)Additionalproteinsareneededfortranscriptioninitiationinvivo:Themediatorcomplex(1)Transcriptionalregulatoryproteins(1)Nucleosome-modifyingenzymes(1’)Tips:1、题目已经问了是真核中的情况，所以必须把相关的真核里的factor都回答出来，不然没分；2、B和C两点非常重要，分值也很大，由此可见把握keypoints的重要性；3、很少同学可以回答出invivo状态下所需要的3种蛋白；4、如果可以答出另外一些细节，比如“TBPbindstoanddistortsDNAusingapsheetinsertedintotheminorgroove”，会有少量加分。Elongation:(7’附+加分)InitiationfactorsarereplacedbyanewsetoffactorsforelongationandRNAprocessing.(或者回答PolIIshedmostofthepreviousfactorsandrecruitothernewfactors)(3’)ThoseelongationfactorsincludeP-TEFbandTFIIS.(如果可以答出factor具体名字，有1分的附加分)RNAprocessingfactorsarealsorecruitedtoPolII.(1’C)ertainfactorsareresponsibleforthecappingofthe5’endoftheRNA(1’)andpolyadenylationofthe3’end,(1)’respectively.Andsplicingfactorscanremoveintrons.(1’)PolIIalsoneedsfactorstodealwithhistonestructures」提到这点会有1分的附加分)Tips:1、不管怎样组织语言回答，一定要体现newfactorREPLACE这层意思，提到replace或相同意思就给3分(由此可见把握keypoints的重要性)；2、P-TEFbandTFIIS相关细节内容可能很多，但不需要回答太多。纵然把相关的detail全部回答出来，也就只算是1个细节，只附加1-2分；3、RNAprocessing相关内容非常重要，有些同学完全没有提到这方面；4、如果回答出相关的其他细节，比如 capping的机制，CPSF和CstF相关因子等，一个细节附加1分。Termination: (7’)Theprocessofterminationcoupleswithpolyadenylation.(1’)Terminationincludestwosteps:mRNArelease;(2’)PolIIdissociation:Therearetwopossiblemodels.Torpedomodel:ThesecondRNAmoleculewithoutacapisrecognizedanddegradedbyapossessiveRNase，thusPolIIisthendissociatedfromDNAtemplate.(2’)Allostericmodel:Thetransferofthe3’processingenzymetoPolIIinducesconformationalchange,thusreducingRolIIprocessivity,andspontaneousterminationoccurs.(2’)Tips:1、要明确的指出mRNA被release掉了，这点很少同学指出；2、很多同学把termination的两个model回答成了Rho-dependent和Rho-independent，这是原核细胞中的机制；3、需要把两个model的具体含义都回答出来，如果仅仅提到名字只给2分。总的Tips:1、一定要把握keypoints，就是答到“点子”上去，比如“promoterescape”，“newfactorreplace”就是非常重要的关键，会占较大比重的分值；2、面对这种大综合题，细节的回答可以为你赢得extrapoints，但是请尽量列举“不同”的细节。如果总是围绕一个细节阐述很多，终究算一个细节，不会得到更多的分数。.Howdoesapre-mRNAintrongetspliced?WhatisthedifferencebetweengroupIIintronsplicingandpre-mRNAintronsplicing?(15’)Howpre-mRNAgetspliced:Step1,formationoftheE(early)complexbyrecognitionofthe5'splicesite,3'splicesiteandAbranchpointbyU1snRNP,U2AFandBBPrespectively.Step2,U2snRNPbindtothebranchsitetoreplaceBBPformsAcomplex.Thebase-pairingbetweenU2snRNAwiththebranchsitemaketheconservedAresidueinthebranchsiteextrudedfromthepairedregion,andthusthisAisreadytocarrythenucleophileattack.Thetri-snRNPU4/U6/U5joinsandAcomplexisarrangedtoBcomplexinwhichthethreesplicesitesarebroughttogether.InthiscomplexU4/U6snRNPsareheldtogethertightlybyextensivebase-pairingbetweenU4andU6snRNAs.Step3,U1leavesthecomplex,andU6occupiesthe5'splicesitebybase-pairing.U4leavesthecomplex,allowingtheRNAcomponentsofU2andU6tobasepairtoproducetheactivesite.ThebranchsiteAattacksthe5'splicesite,formingthe3-wayjunctionandCcomplex.The5'splicesitethenattacksthe3'splicesite,freeingtheintronlariatandformingthemRNAproduct.Difference:thechemistryandtheRNAintermediatesproducedarethesameasthatofthespliceosome-mediatedmRNAsplicing,butthesplicingiscatalyzedbytheintronRNAitself,withouttheaidofproteins..Whatisalternativesplicing?Whyitisimportantforhumanbeing?Howalternativesplicingisregulated?(15’)Alternativesplicing:(4’)Manyprotein-encoded