肺炎链球菌双组分系统WalK蛋白的截短表达及激酶活性分析

摘要

目的 对肺炎链球菌(Streptococcus pneumoniae, s. pn)双组分系统WalK蛋白进行截短表达及激酶活性分析.方法 构建重组表达菌株,通过Ni-NTA亲和层析纯化蛋白,利用Kinase-Glo()化学发光试剂盒检测蛋白活性,并用ELISA和Westernblot方法鉴定其抗原性和抗体特异性.结果 成功获得高浓度、高纯度、有活性的截短WalK重组蛋白.ELISA与WesternBlot检测结果均表明目的蛋白具有良好免疫原性及抗体特异性.结论 成功获得高表达的S. pnWalK截短蛋白,且该蛋白具有较高的激酶活性、免疫原性及抗体特异性,本研究为后续以该蛋白为靶点的疫苗研发及抗菌药物筛选提供实验基础.%Objective To investigate the truncated expression, activity and identification of WalK protein of Streptococcus pneumoniae(S. pn). Methods: The recombinant strain was constructed and the protein was purified by Ni- NTA affinity chromatography. The activity of the protein was measured using the Kinase-Glo kit and the protein was identified by ELISA and Western blot. Results: Successful access to high concentration, high purity,active truncated protein. ELISA and Western Bloting test results show that the successful access to a good immunogenicity and antibody specificity of the target protein. Conclusion: The WalK truncated protein is successfully obtained by highly expressed, and its activity, immunogenicity and antibody specificity are not affected by truncation, which provide the experimental basis for the follow-up vaccine development and antimicrobial drug target research.