直接利用糖质原料产L-丝氨酸谷氨酸棒杆菌glyA基因序列分析  

Analysis of glyA Gene from Corynebacterium glutamicum SYPS-062 Which Can Produce L-serine from Sugar Substances

作　　者：林琳 窦文芳[1] 张晓梅[2] 许泓瑜[2] 许正宏[2] 王正祥[1] 

机构地区：[1]江南大学生物工程学院,无锡214122 [2]江南大学医药学院制药工程实验室,无锡214122

出　　处：《生物技术通报》2008年第5期176-180,共5页Biotechnology Bulletin

基　　金：国家863重点项目(2007AA02Z207);国家973重点项目(2007CB707804);教育部新世纪优秀人才支持计划(NCET-07-0380)

摘　　要：以能够利用糖质原料产L-丝氨酸的谷氨酸棒杆菌(Corynebacterium glutamicum)SYPS-062glyA基因为研究对象,比较C.glutamicum SYPS-062与C.glutamicum模式菌株ATCC13032的glyA基因的异同。分别以SYPS-062及ATCC13032基因组为模板,利用PCR技术获得丝氨酸羟甲基转移酶编码基因glyA。核苷酸序列分析结果表明,来源于SYPS-062和ATCC13032的glyA基因片段全长均为1305bp,编码434个氨基酸,分子量为46.5kD,基因的同源性为99.54%,存在6个核苷酸的差异,引起一个氨基酸残基的突变。将获得的基因分别在大肠杆菌(Escherichia coli)BL21(DE3)中诱导表达。酶活测定结果显示,2种不同菌株来源的重组SHMT的比活力稍有差异,说明SYPS-062glyA基因的差异对其表达产物SHMT蛋白构型及功能影响不大。提示对于C.glutamicum SYPS-062能够利用糖质原料产L-丝氨酸的机制解析应进一步从glyA基因的转录水平、翻译水平及胞内SHMT辅酶的供给情况等方面进行深入研究探讨。Wild corynebacterium glutomicum SYPS-062 can directively product L-serine from sugar substances. This article was forcused on the glyA of SYPS-062 to compare with corynebacterium glutamicum ATCC13032.The two glyA genes were ampilified by PCR method,which used SYPS-062 and ATCC13032 genomes as templates respectively. The result of nucleotide sequence analysis showed that glyA was a 1 305bp fragment,encoding 434 amino acids,with molecular weight of SHMT 46kD and gene homology to 99.54%. Differences exist on six nucleotide which means one amino acid has been changed. The two sources glyA genes were cloned into the vector pET-28a,and then transformed into Escherichia eoli BL21（DE3） to induce expression. It was appeared that the two glyA genes cannot lead to differences of activity of serine hydroxymethyltransferase. Analysis of the mechanism of C. glutamieum SYPS-062 of producing L-serine should be further researched from the glyA gene transcription,translation,as well as the level of intraceUular SHMT coenzyrne supply.

关 键 词：L-丝氨酸 谷氨酸棒杆菌 glyA基因 直接发酵 

分 类 号：Q93[生物学—微生物学]