精氨酸代谢途径抗酸关键基因对乳酸乳球菌Lactococcus lactis NZ9000胁迫抗性的影响  

Influence of key acid-resistant genes in arginine metabolism on stress tolerance in Lactococcus lactis NZ9000

作　　者：张明阳[1,2] 张娟[1,2] 刘龙[1,2] 堵国成[1,3] 陈坚[1,4] 

机构地区：[1]江南大学工业生物技术教育部重点实验室,江苏无锡214122 [2]江南大学生物工程学院,江苏无锡214122 [3]江南大学糖化学与生物技术教育部重点实验室,江苏无锡214122 [4]江南大学粮食发酵工艺与技术国家工程实验室,江苏无锡214122

出　　处：《微生物学通报》2017年第2期314-324,共11页Microbiology China

基　　金：国家自然科学基金项目(No.31470160);国家重点基础研究发展规划项目(973计划)(No.2013CB733902);中国博士后科学基金项目(No.2013M540538;114957);国家高技术研究发展计划项目(863计划)(No.2011AA100901);111计划项目(No.111-2-06)~~

摘　　要：【目的】寻找精氨酸代谢途径中与酸胁迫相关的关键作用因素。【方法】通过在Lactococcus lactis NZ9000中分别过量表达来源于Lactobacillus casei Zhang的精氨酰琥珀酸合成酶(ASS)和精氨酰琥珀酸裂解酶(ASL)改变精氨酸代谢提高酸胁迫抗性。【结果】与对照菌株对比,重组菌株在环境胁迫下表现了较高的生长性能、存活率和发酵性能。生理学分析发现,酸胁迫环境下,重组菌株细胞有较高的胞内NH4+、ATP含量和H+-ATPase活性,并显著提高了精氨酸脱亚胺酶(ADI)途径中的氨基酸浓度。进一步的转录分析发现,天冬氨酸合成、精氨酸代谢相关的基因转录水平上调。【结论】在L.lactis NZ9000中过量表达ASS或ASL可以引发精氨酸代谢流量的上调,进而提高了细胞的多种胁迫抗性。精氨酸合成途径广泛存在于多种微生物中,为微生物,尤其是工业微生物提高胁迫抗性提供了新思路。[Objective] The aim of the present study was to find key factors of arginine metabolism on the stress tolerance. [Methods] Arginine metabolism was switched by argininosuccinate synthase（ASS） or argininosuccinate lyase（ASL） overexpression in L. lactis NZ9000 to enhance the stress tolerance. [Results] The recombinant strains exhibited higher growth performance, viability and fermentation performance compared with the control strain under environmental stresses. Analysis of the physiological data showed that the recombinant cells exhibited higher intracellular p H,intracellular NH4＋ and ATP content, and H＋-ATPase activity under acid stress, and the content of amino acid in arginine deiminase（ADI） pathway was significantly higher than the control strain.Further transcriptional analysis showed that the expression of the genes related to aspartate synthesis and arginine catabolism were up-regulated. [Conclusion] These results suggest that ASS or ASL overexpression in L. lactis NZ9000 can enhance arginine metabolism flux and up-regulated arginine metabolism flux improve the multiple-stress tolerance of cells. As arginine synthesis pathway widely exists in various kinds of microorganisms, results presented in this study provide new idea to enhance stress tolerance of tons of microorganisms especially industrial strains.

关 键 词：LACTOCOCCUS LACTIS 环境胁迫 精氨酰琥珀酸合成酶 精氨酰琥珀酸裂解酶 精氨酸代谢 

分 类 号：Q93[生物学—微生物学]