2013谷氨酸分泌机制及其代谢工程的研究进展_周鹏

，，，（（），214028）：L-谷氨酸是目前全球产量最大的氨基酸。利用发酵法生产谷氨酸需要进行诱导，主要包括：生物素限量；加入表面活性剂；添加青霉素，或使用温度敏感型等。深入揭示谷氨酸生产菌的谷氨酸分泌及其代谢机制，控制优化谷氨酸的分泌，无论从理论还是生产上都有重要意义。另外，随着近年来糖质原料成本的增加，利用代谢工程手段提高现有谷氨酸生产菌糖酸转化率以及开发以木质纤维素为原料的新型谷氨酸代谢途径是研究的热点。以谷氨酸棒杆菌为例，阐述了谷氨酸的分泌机制及其代谢工程的研究进展。：谷氨酸发酵；分泌机制；代谢工程ResearchprogressonsecretionmechanismsandmetabolicengineeringofglutamateoverproductioninCorynebacteriumglutamicumZHOUPeng,XUHong-xian,WANGXin,DUANGang（Genencor（China）Bio-ProductsCo.,Ltd.,Wuxi214028,China）Abstract：Currently,L-glutamateisthelargestaminoacidoftheworld.Glutamateoverproductionbyfermentationisinducedbyspecifictreatmentincludingbiotinlimitation,additionofsurfactantsorpenicillin,usageoftemperaturesensitivemutant.Deeplyclarifyingthemolecularbiologyandmetabolicengineeringstudiesonglutamateproductiontocontrolthesecretionofglutamatehassig-nificantlyimportancebothontheoryandpractice.Furthermore,withtheincreasedcostofrawma-terial,newstudiesarealsofocusedonthemetabolicengineeringofglutamateproductionstrainstoimprovetheproductivityandtoexplorenewpathwayusinglignocellulosesassubstrate.Inthispa-per,therecentprogressinthemolecularmechanismsofglutamateoverproductioninC.glutamicumisreviewedandthecurrentunderstandingofmetabolicengineeringisdiscussed.Keywords：glutamateoverproduction;secretionmechanism;metabolicengineering：，：[e-mail]，（）。1956UdekaKinoshita，（Corynebacteriumglutamicum）L-。，L-，2012250[1]，、，。，70%。发酵科技通讯424201310—19—发酵科技通讯4250，，。，。，，，，，，[2]，，，。，。1。，，。1.11968，Takinami[3]，，，，，，。（leakmodel），，，H+,K+，Cl-，[4-6]，。1.2，[7]，1。。（1）（ODHC）；（2）；（3）。1.2.1ODHC活性的降低，3，2-(E1o)，（E2o），（E3）。，，ODHC，Kawahara[8]，，ODHC，ODHC，E1oodhA，，[9]。ODHCOdhI/PknGPpp。OdhI。odhICFHAodhA，ODHC。PknGodhIN1—20—，：4Thr14，，FHA，odhIODHC。PppOdhI，Ppp，odhI[10-12]。Belanger[13]GarA，odhI，Villarino[14]GarAPknB。PknBPknG/。，（、、），。Shimizu[15]α-（ICDH）,（GDH）ODHC，（2）。icdhgdh，3，70%TCA。ODHC52%，75%，ODHC，。1.2.2谷氨酸运输载体，Nakamura[16]，NCgl1221，。，NCgl1221，C，NYggB[17]。Berrier[18]，。Hashimoto[19]NCgl1221。、40、，NCgl1221，NCgl1221，，。，NCgl1221，NCgl1221。1.2.3细胞膜状态的改变、。CoA。。，，。，。Kimura[20]C.glutamicumTween40DtsR1，DtsR1，DtsR2，CoAβ。Dt-sR1AccBC，J覿ger[21]AccBC，2ICDH,GDH,ODHC—21—AccBCCoAα，Dt-sR1DtsR2β，DtsR1/AccBCDtsR2/AccBC，。，dtsR1，[22]。Hirasawa[23]ltsA，ltsA，，。ltsA、。。2，，[24]。，，。，。2.1，odhA、DtsR1，odhIODHC。，，81.7%，CoACO2，。（PKT）CO2，-6-4，5-5-3，（PTA）CoA，（3）。CoACO2，56，81.7%98%。Chinen[25]PKT，60%69%。ACO2，BPKTCO23发酵科技通讯42—22—2.2，。，，。、。，，（4）。Kawaguchi[26]xylAxylB5-，3-；AraA，AraB，5-AraD5-，，[27]。Kotrba[28]C.glutamicumR，bglF，，-β-BglA。Tsuchidate[29]，，。，。C.glutamicum。Rittmann[30]glpFDKC.glu-tamicum，，。AraA-；AraB-；AraD-5--；AraE-；Tpi-；XylA-；XylB-；BglF-PTS；BglA--β-；CelE-；IolT-；PtsG-PTS；GlpF-；GlpK-；GlpD-3-4，：4—23—发酵科技通讯423，，，50，。，，。，，。。，，。，、，，[31]。：[1]BeckerJ,WittmannC.Systemsandsyntheticmetabolicengi-neeringforaminoacidproduction-theheartbeatofindustrialstraindevelopment[J].CurrentOpinioninBiotechnology,2012,23(5):718-726.[2]，，.[J].，2008,27(7):983-989.[3]TakinamiK,YoshiiH,YamadaY,etal.ControlofL-glutamicacidfermentationbybiotinandfattyacids[J].AminoAcidandNucleicAcid,1968,18:120-126.[4]GutmannM,HoischenC.Carrier-mediatedglutamatesecretionbyCorynebacteriumglutamicumunderbiotinlimitation[J].BiochimicaetBiophysicaActa,1992,1112:115-123.[5]HoischenC,Kr覿merR.MembranealterationisnecessarybutnotsufficientforeffectiveglutamatesecretionbyCorynebacteri-umglutamicum[J].JournalofBacteriology,1990,172:3409-3416.[6]NeubeckM,PrennerE,HorvartP,etal.Membranefluidityinglutamicacid-producingbacteriaBrevibacteriumsp.ATCC13869[J].ArchivesofMicrobiology,1993,160:101-107.[7]EggelingL,BottM.HandbookofCorynebacteriumglutamicum[M].CRCPress,2005.444-457.[8]KawaharaY,TakahashiFK,ShimizuE,etal.Relationshipbe-tweentheglutamateproductionandtheactivityof2-oxoglu-taratedehydrogenaseinBrevibacteriumlactofermentum[J].Bio-scienceBiotechnologyandBiochemistry,1997,61:1109-1112.[9]UsudaY,TujimotoN,AbeC,etal.MolecularcloningoftheCorynebacteriumglutamicum(‘Brevibacteriumlactofermentum’AJ12036)odhAgeneencodinganoveltypeof2-oxoglutaratedehydrogenase[J].Microbiology,1996,142:3347-3354.[10]NiebischA,KabusA,SchultsC,etal.CorynebacterialproteinkinaseGcontrols2-oxoglutaratedehydrogenaseactivityviaphosphorylationstatusoftheOdhIprotein[J].BiologicalChem-istry,2006,281:12300-12307.[11]SchultzC,NiebischA,GebelL,etal.GlutamateproductionbyCorynebacteriumglutamicum:dependenceontheoxoglutaratedehydrogenaseinhibitorproteinOdhIandproteinkinasePknG[J].AppliedMicrobiologyandBiotechnology,2007,76:691-700.[12]SchultzC,NiebischA,SchwaigerA,etal.Geneticandbio-chemicalanalysisoftheserine/threonineproteinkinasesPknA,PknB,PknGandPknLofCorynebacteriumglutamicum:evi-dencefornon-essentialityandforphosphorylationofOdhIandFtsZbymultiplekinases[J].MolecularMicrobiology,2009,74:724-741.[13]BelangerAE，HatfullGF.Exponential-phaseglycogenrecy-clingisessentialforgrowthofMycobacteriumsmegmatis[J].JournalofBacteriology,1999,181:6670-6678.[14]VillarinoA,DuranR,WehenkelA,etal.ProteomicidentificationofM.tuberculosisproteinkinasesubstrates:PknBrecruitsGarA,aFHAdomaincontainingprotein,throughactivationloop-mediatedinteractions[J].J