通过定点突变提高乳链菌肽对热及pH的稳定性  

Improving heat and pH stability of nisin by site-directed mutagenesis

作　　者：路遥[1] 蒋立科[1] 陈美玲[2] 还连栋[2] 钟瑾[2] 

机构地区：[1]安徽农业大学生命科学学院,合肥230036 [2]中国科学院微生物研究所,北京100101

出　　处：《微生物学报》2010年第11期1481-1487,共7页Acta Microbiologica Sinica

基　　金：国家"863计划"(2006AA10Z319);中国科学院知识创新重要方向项目(KSCX2-YW-G-016)~~

摘　　要：【目的】通过定点突变技术改变乳链菌肽(nisin)特定位置氨基酸,获得性质改善的nisin突变体,为扩大其应用范围提供依据。【方法】在抑菌谱扩大的nisin单突变体M21K nisinZ的基础上,对M21K nisZ基因第29位丝氨酸密码子进行定点突变;将其克隆至乳酸菌表达载体pMG36e,并在Lactococcus lactis NZ9800中进行表达;双突变体M21K/S29K nisinZ经分离纯化后检测其在抑菌活性、抑菌谱和稳定性等方面的变化。【结果】与单突变体M21K nisinZ及野生型nisinZ(wild-type,WT)相比,双突变体M21K/S29K nisinZ对指示菌的抑菌活性虽有所下降,但其对温度及pH值的稳定性有显著提高。同时其抑菌谱与M21K nisinZ相同,可抑制革兰氏阴性菌,扩大了WT的抑菌谱。【结论】通过改变nisin分子特定位置的氨基酸可以改善nisin分子的理化性质,有可能得到应用范围更广的nisin品种。[Objective]The aim of this study was to optimize the property of nisin through altering its specific amino acid by site-directed mutagenesis method. [Methods]On the basis of M21K nisinZ,a former reported nisinZ mutant that exhibited antimicrobial activity against Gram-negative bacteria,the 29th amino acid of it was mutated from serine to lysine. The mutant M21K /S29K nisZ gene was cloned into vector pMG36e and the recombinant plasmid was introduced into Lacotococcus lactis NZ9800. The resulting M21K /S29K nisinZ was then isolated and purified,and its antibacterial activity,antibacterial spectrum and stability were analyzed and compared to those of M21K nisinZ and nisinZ. [Results] Compared with wild-type nisinZ and M21K nisinZ,the M21K /S29K nisinZ displayed reduced antimicrobial activity,but showed significantly increased stabilities to heat and pH stress. Moreover,M21K /S29K nisinZ also exhibited antimicrobial activity against Gram-negative bacteria as M21K nisinZ did. [Conclusion]By changing the 29^th amino acid of nisin,we can optimize the property of nisin,especially its stability to heat and pH stress.

关 键 词：乳链菌肽 稳定性 温度 PH 定点突变 

分 类 号：Q933[生物学—微生物学]