Purification and properties of S-hydroxymethylglutathione dehydrogenase of Paecilomyces variotii no. 5, a formaldehyde-degrading fungus 英文参考文献.docVIP

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Purification and properties of S-hydroxymethylglutathione dehydrogenase of Paecilomyces variotii no. 5, a formaldehyde-degrading fungus 英文参考文献.doc

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Purification and properties of S-hydroxymethylglutathione dehydrogenase of Paecilomyces variotii no. 5, a formaldehyde-degrading fungus 英文参考文献

Fukudaetal.AMBExpress2012,2:32 /content/2/1/32 ORIGINAL ARTICLE OpenAccess Purificationandpropertiesof S-hydroxymethylglutathionedehydrogenaseof Paecilomycesvariotiino.5,a formaldehyde-degradingfungus RyoheiFukuda,KazuhiroNagahama,KohsaiFukuda,KeisukeEkino,TakujiOkaandYoshiyukiNomura* Abstract S-hydroxymethylglutathionedehydrogenasefromPaecilomycesvariotiiNo.5strain(NBRC109023),isolatedasa formaldehyde-degradingfungus,waspurifiedbyaprocedurethatincludedammoniumsulfateprecipitation,DEAE- Sepharoseandhydroxyapatitechromatographyandisoelectrofocusing.Approximately122-foldpurificationwas achievedwithayieldof10.5%.Theenzymepreparationwashomogeneousasjudgedbysodiumdodecyl polyacrylamidegelelectrophoresis(SDS).Themolecularmassofthepurifiedenzymewasestimatedtobe 49kDabySDSandgelfiltration,suggestingthatitisamonomer.EnzymeactivitywasoptimalatpH8.0and wasstableintherangeofpH7.0–10.Theoptimumtemperatureforactivitywas40°Candtheenzymewasstable upto40°C.TheisoelectricpointwaspH5.8.Substratespecificitywasveryhighforformaldehyde.Besides formaldehyde,theonlyaldehydeoralcoholtestedthatservedasasubstratewaspyruvaldehyde.Enzymeactivity wasenhancedbyseveraldivalentcationssuchasMn2+(179%),Ba2+(132%),andCa2+(112%)butwascompletely inhibitedbyNi2+,Fe3+,Hg2+,p-chloromercuribenzoate(PCMB)andcuprizone.Inactivationoftheenzymeby sulfhydrylreagents(Hg2+andPCMB)indicatedthatthesulfhydrylgroupoftheenzymeisessentialforcatalytic activity. Keywords:S-hydroxymethylglutathionedehydrogenase,Enzymepurification,Formaldehydemetabolism, Paecilomycesvariotii,SHenzyme Introduction experimental specimens. Formaldehyde is a highly toxic Formaldehyde is a ubiquitous compound that is a prod- compound due to nonspecific reactivity with proteins uctofbiologicalsources(fromphotooxidationofatmos- andnucleicacids (Grafstromet al. 1983), soit isan en- pheric hydrocarbons) (Levy 1971; Zimmerman et al. vironmentalpollutant. 1978)andenvironmentalsources(emissionsfromindus- Toaddresstheproblemofformaldehydepollu