乳酸乳球菌acmAas基因的克隆与基因中断突变株特性研究  

Cloning and Characterization of acmAas from Lactococcus lactis and Effect of a Truncated Derivative on Daughter-Cell Segregation and Growth

作　　者：张震[1] 李小华[1] 黄新凤[1] 邵小虎[1] 李林[1] 

机构地区：[1]华中农业大学农业微生物学国家重点实验室,武汉430070

出　　处：《华中农业大学学报》2009年第5期577-582,共6页Journal of Huazhong Agricultural University

基　　金：国家自然科学基金项目(3067005430370026)资助

摘　　要：克隆了乳酸乳球菌(Lactococcus lactis)菌株AS1.2829的1种主要细胞壁自溶素-N-乙酰葡萄糖胺糖苷酶的编码基因acmAas,并对其编码蛋白AcmAas的结构进行了预测,发现该蛋白由1个具催化活性的N-末端和1个具细胞壁锚定活性的C-末端结构域组成,其中C-末端又由3个LysM(lysin motif)结构域组成,具有典型的"βαβα"型二级结构。进一步通过体外在acmA基因中引入红霉素抗性基因(Emr)而构建具有与宿主菌染色体acmA基因进行整合作用的重组基因,并导入宿主菌后筛选得到发生染色体整合重组的重组菌株MB257,对该菌株的形态与生长特性进行了初步研究,结果表明该菌株所携带的acmAasN-lysM1基因在保留其N端结构域和1个lysM编码区后,其编码产物可发挥其完整生物学活性。A gene encoding a major peptidoglycan hydrolase（autolysin） of Lactococcus lactis, N-acetylglucosaminidase（AcmA） was cloned from L. lactis wild-type strain AS1. 2829 and the putative structure and domain organization of the encoded protein, AcmAas,was characterized. The overall amino acid sequence of AcmAas was structurally distinguished as an N-terminal catalytic domain with a 57 aa of signal peptide sequence and a C-terminal cell-wall binding domain with three tandemly aligned lysin motifs（LysM） that constitutes a modular secondary structure of ＂βααβ＂, and a further blast analysis revealed a significant sequence similarity subjected to the previously identified N-acetylglucosaminidase of L. lactis subsp, lactis IL1403, suggesting AcmAas could be an N-acetylglucosaminidase of AS1. 2829. To investigate whether the sole LysM domain is functional for binding to the cell-wall, a chromosome-inte- grated engineered strain of AS1. 2829, MB257, was constructed by introducing a chimeric gene of ＂acmAasC1-Ern-acmAasC2＂,which elicited an intracellular dual exchanges between host chromosome and the introduced gene. Results showed that MB257 strain exhibited normal daughter-cell segregation and growth pattern,indicating that the sole LysM domain can serve as the functional active domain for cellwall binding.

关 键 词：乳酸乳球菌 acmA 克隆 染色体整合重组 

分 类 号：Q939.9[生物学—微生物学]