结核分枝杆菌RD1区重组11 kD蛋白的克隆表达及效力

• 作 者: 陈保文 徐苗 徐敬华 沈小兵 苏城 王国治
• 机 构: 中国药品生物制品检定所,中国药品生物制品检定所,中国药品生物制品检定所,中国药品生物制品检定所,中国药品生物制品检定所,中国药品生物制品检定所 北京100050,北京100050,北京100050,北
• 出 处: 中国生物制品学杂志
• 发表日期: 2006年05期
• 关键词: 结核分枝杆菌 重组11kD蛋白 效力

目的在大肠杆菌中表达结核分枝杆菌RD1区11kD蛋白,并检测表达产物的效力。方法构建11kD蛋白编码基因原核表达载体,转化大肠杆菌BL21(DE3)IPTG诱导表达,离子层析柱分离纯化重组蛋白。以结核分枝杆菌致敏豚鼠进行皮肤变态反应(DTH)测定。结果结核分枝杆菌11kD蛋白在大肠杆菌中以可溶形式表达,表达蛋白占总菌体蛋白的33%以上,纯化后纯度达95%以上,并可诱导结核分枝杆菌致敏豚鼠产生迟发型超敏反应,5μg/ml11kD蛋白的效力与50IU/ml TB-PPD相当。结论重组11kD蛋白已在大肠杆菌中成功表达,并有望成为结核病皮试鉴别诊断用新试剂。

Objective To express 11kD protein of Mycobacterium tuberculosis RD1 region in Escherichia coli and test the efficacy of the expressed product. Methods The prokaryotic expression vector encoding the 11kD gene was constructed and transformed into E. coli BL21 (DE3) for expression induced by IPTG. The recombinant protein was isolated and purified by ion chromatography. Skin allergy (DTH) determination was performed on M. tuberculosis sensitized guinea pigs. Results Mycobacterium tuberculosis 11kD protein was expressed in soluble form in E. coli. The expressed protein accounted for more than 33% of the total bacterial proteins. The purity of the 11kD protein was over 95% after purification, and induced the delayed type of Mycobacterium tuberculosis sensitized guinea pigs Hypersensitivity, 5μg / ml 11kD protein potency and 50IU / ml TB-PPD equivalent. Conclusion The recombinant 11 kD protein has been successfully expressed in E. coli and is expected to become a new reagent for the differential diagnosis of tuberculosis skin test.