为什么纯化酶 Why Purify Enzymes by Arthur Kornberg.pdf

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1、CHAPTERONEWhyPurifyEnzymes?wArthurKornberg‘‘Don’twastecleanthinkingondirtyenzymes’’isanadmonitionofEfraimRacker’swhichisatthecoreofenzymologyandgoodchemicalpractice.Itsimplysaysthatdetailedstudiesofhowanenzymecatalyzestheconversionofonesubstancetoanotherisgenerallyawasteoftime

2、untiltheenzymehasbeenpurifiedawayfromtheotherenzymesandsubstancesthatmakeupacrudecellextract.Themixtureofthousandsofdifferentenzymesreleasedfromadisruptedliver,yeast,orbacterialcelllikelycontainsseveralthatdirectotherrearrangementofthestartingmaterialandtheproductoftheparticul

3、arenzyme’saction.OnlywhenwehavepurifiedtheenzymetothepointthatnootherenzymescanbedetectedcanwefellassuredthatasingletypeofenzymemoleculedirectstheconversionofsubstanceAtosubstanceB,anddoesnothingmore.Onlythencanwelearnhowtheenzymedoesitswork.Therewardsforthelaborofpurifyingane

4、nzymewerelaidoutinaseriesofinspirationalpapersbyOttoWarburginthe1930s.FromhislaboratoryinBerlin-Dahlemcamethedisciplineandmanyofthemethodsofpurifyingenzymesandwiththosetheclarificationofkeyreactionsandvitaminfunc-tionsinrespirationandthefermentationofglucose.Warburg’scontribut

5、ionsstrengthenedtheclassicapproachtoenzymologyinauguratedwithEduardBu¨chner’saccidentaldiscovery,attheturnofthiscentury,ofcell-freeconversionofsucrosetoethanol.Onetracksthemolecularbasisofcellularfunction—alcoholicfermentationinyeast,glycolysisinmuscle,lumines-cenceinafly,orth

6、ereplicationofDNA—byfirstobservingthephenome-noninacell-freesystem.Thenoneisolatestheresponsibleenzyme(orenzymes)byfractionationofthecellextractandpurifiesittohomogene-ity.Thenonehopestolearnenoughaboutthestructureoftheenzymetoexplainhowitperformsitscatalyticfunctions,responds

7、toregulatorysignals,andisassociatedwithotherenzymesandstructuresinthecell.Byareverseapproach,callitneoclassical,especiallypopularinrecentdecades,onefirstobtainsastructureandthenlooksforitsfunction.Theproteinispreferablysmallandstable,andhasbeenamplifiedbycloningoriscommerciall

8、yavailable.Byintensivestudyoftheproteinandhomologousproteins,