摘要
目的:克隆、表达肺炎链球菌LicC及LicC3′端25个氨基酸缺失片段(记作△LicC)基因;分析、比较目标蛋白活性,证实LicC部分缺失对其活性的影响。方法:通过基因重组技术构建重组质粒pQE80-licC、pQE80-△licC,转化大肠杆菌BL21,异丙基-β-D-硫代半乳糖苷(IPTG)诱导表达,自建生物荧光技术测定酶活性。结果:成功构建重组表达载体,获得可溶性LicC和LicC截短体蛋白,活性测定表明LicC截短体活性显著低于LicC(P<0.05)。结论:自建生物荧光技术测定LicC活性准确、可靠;证实LicC3′端25个氨基酸对其活性有重要影响;提示抑制LicC活性有可能成为一种有效的杀灭耐药肺炎链球菌的措施。
AIM：To clone and express licC and its truncated form genes（25 amino acids from 3＇-terminus were deleted and named △LicC）of Streptococcus pneumoniae,analyze the enzymatic activities of the proteins. METHODS：The recombinant plasmid pQE80-licC,pQE80-△licC were constructed,and the target proteins were expressed in E.coli BL21 under isopropy-β-D-thiogalactoside（IPTG）induction. The proteins＇ activities were determined using bioluminescence test based on firefly luciferase assay system. RESULTS：The prokaryotic expression vector pQE80-licC,pQE80-△licC were successfully constructed and identified.The soluble proteins were obtained through inducing expression in E.coli BL21.It was showed that the activity of △LicC was markedly lower than that of LicC （P〈0.05）. CONCLUSION：The homemade bioluminescence assay method can miner the activity of LicC reliably and accurately. The important role of 25 amino acids from 3＇-terminus for the activity of LicC was confirmed,and it was suggested that suppressing of LicC maybe was a useful method for treatment S.pneumococcal infection,especially drug resistant strain.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2010年第4期337-339,343,共4页
Chinese Journal of Cellular and Molecular Immunology
基金
国家自然科学基金资助项目(30873074)
