CTX-M-3型超广谱β-内酰胺酶原核表达条件的研究  

A Study on Prokaryotic Expression of An Extended-spectrum β-lactamase CTX-M-3

作　　者：徐韫健[1] 李翊泉[2] 廖伟娇[1] 江洁华[1] 张东梅[1] 

机构地区：[1]广州医学院第一附属医院检验科,广州510120 [2]广州医学院第一附属医院输血科,广州510120

出　　处：《热带医学杂志》2010年第2期123-125,129,共4页Journal of Tropical Medicine

基　　金：广州市医药卫生科技(No.2007-YB-156);广州市医药卫生科技(No.2008-YB-149)

摘　　要：目的探讨重组质粒pET-28a(+)/CTX-M-3分别在大肠埃希菌ER2566和BL21(DE3)中进行原核表达的最佳条件。方法分别在不同的宿主菌、诱导温度、诱导时间和诱导剂浓度中诱导pET-28a(+)/CTX-M-3融合表达载体,目的产物经SDS-PAGE和BandScan凝胶分析软件分析,以获得最佳表达条件。结果表达载体的最佳诱导条件是重组质粒在BL21(DE3)中18℃诱导24h,IPTG终浓度为0.8mmol/L;表达载体在最佳条件下表达时,目的蛋白的最高表达量占菌体总蛋白的33%。结论获得pET-28a(+)/CTX-M-3在大肠埃希菌中表达CTX-M-3型超广谱β-内酰胺酶(extended-spectrum β-lactamase,ESBLs)蛋白的最佳条件,为此酶的大量纯化奠定了基础。Objective To optimize the expression conditions of pET-28a（＋）/CTX-M-3 in E.coli ER2566 and BL21 （DE3）. Methods pET-28a （＋）/CTX-M-3 expression vector was expressed in different bacteria host. The optimum conditions for expression, such as the induction temperatures, induction time and the concentration of inducer were determined.The protein expression level was evaluated using SDS-PAGE and BandScan gel analysis software. Results The optimal induction conditions for the expression in E.coli BL21 （DE3） were 18℃, 24 h of incubation and 0.8 mmol/L of IPTG.Under these conditions, the maximum amount of the recombinant protein expressed was 33% of the total cellular protein.Conclusion The optimum conditions for the expression of CTX-M-3 in E.coli were determined. The conditions can be used for the future large scale production of CTX-M-3.

关 键 词：CTX—M一3 超广谱β一内酰胺酶 原核表达 条件 

分 类 号：R392[医药卫生—免疫学]