地衣芽胞杆菌乙醛酸循环对聚谷氨酸合成的影响  

Effect of glyoxylate cycle in Bacillus licheniformis on the synthesis of poly glutamic acid

作　　者：田光明[1] 游蕾[2] 陈守文[3,4] 

机构地区：[1]长江大学湿地生态与农业利用教育部工程研究中心动物科学学院,湖北荆州434020 [2]长江大学城市建设学院,湖北荆州434023 [3]湖北大学生命科学学院,湖北武汉4300623 [4]华中农业大学生命科学技术学院,湖北武汉430062

出　　处：《生物技术》2017年第1期78-84,共7页Biotechnology

基　　金：湖北省教育厅科学技术研究计划青年人才项目("应激蛋白(HSP70)对黄鳝应激反应的调控作用研究";No.q20151306);湿地生态与农业利用教育部工程研究中心开放课题("饲料中花生四烯酸对黄鳝抗应激能力的影响";No.KF201509)

摘　　要：[目的]了解乙醛酸循环在地衣芽胞杆菌WX-02生物合成聚谷氨酸中作用,为聚谷氨酸生产提供新的解决方法。[方法]采用基因工程手段,以地衣芽胞杆菌WX-02为原始菌株,分别增强表达和敲除异柠檬酸裂解酶ace A基因,检测发酵过程中聚谷氨酸产量、生物量、胞内外代谢物和相关基因转录量。[结果]增强表达异柠檬酸裂解酶ace A基因后,胞内谷氨酸浓度显著升高(483.42 ng/m L/Log(CFU)),溢流代谢产物减少(乙酸5.41 g/L、乙偶姻5.82 g/L、2,3-丁二醇7.31 g/L),聚谷氨酸生物合成产量为11.74 g/L,相比原始菌株提高15%。谷氨酸脱氢酶roc G基因、谷氨酸消旋酶glr基因和聚谷氨酸合成酶复合体中pgs B基因转录水平相对原始菌株分别提高1.61倍、1.32倍和1.24倍。[结论]增强乙醛酸循环可以降低地衣芽胞杆菌WX-02乙酸等溢流代谢产物合成,提高胞内谷氨酸合成能力,并上调聚谷氨酸合成酶基因转录水平,最终提高聚谷氨酸生物合成产量。[Objective]The objective of this work was to investigate the effects of glyoxylate cycle on polyglutamic acid（PGA）production of Bacillus licheniforims WX-02 and solve the problem in PGA production. [Methods]The yield of PGA,biomass,intracellular metabolite,and transcription levels of genes involved in PGA synthesis were measured by enhancing and knocking out the isocitratelyase encoded by ace A gene in Bacillus licheniformis WX-02,respectively. [Results]By enhanced expression of ace A gene in vivo,the concentration of intracellular glutamic acid was improved（483. 42 ng/m L/Log（CFU））,and the overflow products of acetic acid,acetoin and 2,3-butanediol were decreased to 5. 41 g/L,5. 82 g/L,and 7. 31 g/L,respectively.Moreover,the yield of PGA reached 11. 74 g/L,15% higher than the native strain. Gene transcript levels of glutamate dehydrogenase encoded by roc G gene,glutamate racemase encoded by glr gene and pgs B gene in polyglutamate synthetase were confirmed to be improved by 1. 61-fold,1. 32-fold,and 1. 24-fold over that of the control. [Conclusion]Decrease of overflow metabolites synthesis,increase of the intracellular glutamate synthesis,and upregulation of gene transcript levels of poly glutamate synthetase were confirmed in PGA biosynthesis,which might be the reason for the increased PGA yield.

关 键 词：地衣芽胞杆菌 乙醛酸循环 聚谷氨酸 基因转录 发酵 

分 类 号：Q939.97[生物学—微生物学]