幽门螺杆菌omp22基因工程菌蛋白表达条件优化、纯化及活性鉴定  

OPTIMIZATION OF EXPRESSION CONDITIONS FOR OUTER MEMBRANE PROTEIN OMP22 GENE OF HELICOBACTER PYLORI IN E.COLI AND IDENTIFICATION OF PURIFIED PROTEIN IMMUNOCOMPETENCE

作　　者：黄学勇[1] 段广才[2,3] 范清堂[3] 郗园林[2] 

机构地区：[1]河南省疾病预防控制中心,郑州450016 [2]郑州大学公共卫生学院流行病学教研室 [3]河南省分子医学重点学科开放实验室

出　　处：《现代预防医学》2010年第8期1507-1509,共3页Modern Preventive Medicine

基　　金：河南省医学创新人才基金资助项目(2000-84);河南省科技厅科技攻关项目基金资助课题(0424410035)

摘　　要：[目的]优化幽门螺杆菌omp22基因工程菌(pMAL-c2X-omp22-TB1)的表达条件,并对目的蛋白进行纯化和免疫活性鉴定。[方法]在不同诱导时机、诱导剂(IPTG)浓度、诱导时间下诱导pMAL-c2X-omp22-TB1的表达,测定目的蛋白的表达量。在优化条件下诱导工程菌表达,采用SDS-PAGE方法对表达产物分析;Amyloss树脂预装柱对可溶性OMP22蛋白进行分离、纯化,对纯化蛋白做Western blot分析。[结果]工程菌培养2h时加入IPTG(终浓度为0.3mmol/L)诱导4h,目的蛋白表达量达到菌体总蛋白的25%以上;经纯化得到了较高纯度(﹥90%)的目的蛋白,并具有良好的免疫原性。[结论]建立了从TB1(pMAL-c2X-omp22)可溶性表达产物中纯化较高纯度OMP22融合蛋白的方法,为进一步的动物实验和诊断性抗原的研制以及工程菌高效表达生产工艺的研究打下了基础。[Objective] To optimize the expressing conditions of gene engineering E.coli strain pMAL-c2X-omp22-TB1 and to purify its expressing products,which immunogenicity were identified. [Methods] pMAL-c2X-omp22-TB1 was induced by different contents of IPTG for different inducing time at different inducing opportunity. The expression products were analyzed by using the SDS-PAGE method; Soluble OMP22 protein was purified by Amyloss pre-packed column and then the purity was detected with Western blot. [Results] The gene engineering E.coli strain growing two hours was induced for four hours by 0.3mmol / L IPTG,the expression amounts of fusion protein took account for 25% of the total bacterium protein,and purified OMP22 protein was confirmed to more than 90%. OMP22 had a high immunocompetence. [Conclusion] An effective method for purifying OMP22 protein of Helicobacter pylori from soluble offspring was established,which will be helpful for animal study and producing OMP22 protein.

关 键 词：幽门螺杆菌 基因工程 高效表达 纯化 

分 类 号：R339.5[医药卫生—人体生理学]