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1、DirectedMolecularEvolutionofProteins:orHowtoImproveEnzymesforBiocatalysis.EditedbySusanneBrakmannandKaiJohnssonCopyrightã2002Wiley-VCHVerlagGmbH&Co.KGaAISBNs:3-527-30423-1(Hardback);3-527-60064-7(Electronic)4ConstructionofEnvironmentalLibrariesforFunct
2、ionalScreeningofEnzymeActivityRolfDanielAssemblagesofenvironmentalmicroorganismsoftenencompassabewilderingarrayofphysiological,metabolic,andgeneticdiversity.Althoughinvisibletothenakedeye,microorganismsdominatethebiosphere[1].Forexample,ithasbeencalcul
3、atedthat1gofsoilmaycontainupto4000differentbacterialspecies[2].Currentestimatesofthetotalnumberofmicrobialspeciesonearthrangefrom1,000,000to100,000,000,butonlyapproximately5000microbialspeciesaredescribedintheliterature[3,4].Inaddition,therecentresearc
4、hinmolecularmicrobialecologyprovidescompellingevi-dencefortheexistenceofmanynoveltypesofmicroorganismsintheenvironmentinnumbersandvarietiesthatdwarfthoseofthecomparativelyfewmicroorganismsstu-diedtodate.TheseresultswereobtainedbyestimationofDNAcomplexi
5、tyanddis-coveryofmanyunique16SrRNAgenesequencesfromvariousenvironmentalsources[5,6].Thus,theentiretyofthemicrobialgenomesfoundinnature,termedthemetagenomebysomeauthors[7],harborsvastlymoregeneticinformationthaniscontainedinthestudiedandculturedsubset.C
6、orrespondingly,thepotentialofmicro-organismsandtheirenzymesinthesynthesisofnewcompoundsisbynomeansexhausted.Theassessmentandexploitationofthisenormousmicrobialdiversityof-fersanalmostunlimitedpoolofnewgenes,geneproducts,andbiosyntheticpathwaysforbiotec
7、hnologyandotherpurposes.Theclassicalandcumbersomeapproachtoisolatenewenzymesfromenvironmen-talsamplesisenrichmentandscreeningofawidevarietyofmicroorganismsforthedesiredactivity.Theenzymesandthecorrespondinggenesarethenrecoveredfromtheidentifiedorganism
8、s.Traditionalcultivationtechniquesrequirethatthedifferentmicroorganismsderivedfromanenvironmentalsamplearecultivatedonanappro-priategrowthmediumandthenseparateduntilindividualclonesareisolated.Inthisway,alargefractionofthemicrobialdiver
