一种筛选高产(+/-)-γ-内酰胺酶菌株方法的建立  

Establishment of a Method for Screening High Yield(+/-)-γ-lactamase Producing Strains

作　　者：王金昌[1] 王建军[2] 关丽梅[1] 占智高[1] 靳亮[1] WANG Jinchang;WANG Jianjun;GUAN Limei;ZHAN Zhigao;JIN Liang(Institute of Microbiology,Jiangxi Academy of Sciences,330096,Nanchang,PRC;Institute of Microbiology Chinese Academy of Sciences,100101,Beijing,PRC)

机构地区：[1]江西省科学院微生物研究所,南昌330096 [2]中国科学院微生物研究所,北京100101

出　　处：《江西科学》2018年第6期919-922,941,共5页Jiangxi Science

基　　金：江西省科学院对外合作重点项目(2013H006)

摘　　要：(+)-γ-内酰胺酶能够拆分(+/-)-γ-内酰胺得到(-)-γ-内酰胺,(-)-γ-内酰胺酶能够拆分(+/-)-γ-内酰胺得到(+)-γ-内酰胺。目前只有几种具有(+/-)-γ-内酰胺酶的微生物被报道,都是用N-乙酰-L-苯丙氨酸做为唯一碳源从土壤里筛选能利用N-乙酰-L-苯丙氨酸的菌株,它们有可能具有(+/-)-γ-内酰胺酶,再用手性高效液相色谱分析法复筛。但是这种筛选方法有点盲目性,工作量大。本实验在此基础上,先利用经典的复制平板和茚三酮染色技术,再用氨基酸显色法复筛得到3株阳性菌株,最后用手性高效液相色谱分析法确认。结果筛选到的3号菌株具有(+)-γ-内酰胺酶功能,ee值达到100%,22号和29号菌株具有(-)-γ-内酰胺酶功能,ee值达到50%。此种筛选方法的建立,以后也可用于类似的易溶,又不显色底物的降解菌株和阳性克隆子的筛选。The（ ＋ ）-γ-lactamases can resolute （ ＋/- ）-γ-lactam and produce （ - ）-γ-lactam. The （ - ） -γ-lactamases can resolute （ ＋ / - ） -γ-lactam and produce （ ＋ ） -γ-lactam. At present, only a few microorganisms with（ ＋/- ）-γ-lactamase were reported to be screened from the soil using the N-acetyl-L-phenylalanine as the sole source of carbon,they may have（ ＋/- ）-γ-lactamase and then they are rescrened by chiral high performance liquid chromatography. But this method is a bit blind- ness and heavy workload. On the basis of them, We preliminarily screened many positive strains combining with the classic duplicating plate and ninhydrin dyeing technology, and rescreened three positive strains by color analysis of amino acids, finally by chiral high performance liquid chromatography analysis. Results：strain 3 had the function of （ ＋ ）-γ-lactamase, the EE value reached 100% , and the 22 and 29 strains had the function of（ - ）-γ-lactamase,and the EE value reached 50%. The establishment of this screening method can be used to select the degradation strains and the positiveclones of soluble substrate where a selective staining method exists in the future.

关 键 词：(+/-)-γ-内酰胺酶 γ-内酰胺 生物催化 

分 类 号：Q936[生物学—微生物学]