Expression and purification of Mycobacterium tuberculosis ESAT-6 and MPT64 fusion protein and its immunoprophylactic potential in mouse model

• Published in: Protein expression and purification Vol. 59; no. 2; pp. 189 - 196
• Main Authors: Bai, Yinlan, Xue, Ying, Gao, Hui, Wang, Limei, Ding, Tianbing, Bai, Wentao, Fan, Ailin, Zhang, Jianfang, An, Qunxing, Xu, Zhikai
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.06.2008
• Subjects: Animals; Antigens, Bacterial - biosynthesis; Antigens, Bacterial - isolation & purification; Antigens, Bacterial - therapeutic use; Bacterial Proteins - biosynthesis; Bacterial Proteins - isolation & purification; Bacterial Proteins - therapeutic use; Blotting, Western; Chromatography, Affinity; Cloning, Molecular; Disease Models, Animal; ESAT-6; Escherichia coli; Escherichia coli - genetics; Expression; Genetic Vectors; Humans; Interferon-gamma - metabolism; Interleukin-12 - metabolism; Lymphocyte Activation; Mice; Mice, Inbred C57BL; MPT64; Mycobacterium tuberculosis; Mycobacterium tuberculosis - immunology; Mycobacterium tuberculosis - isolation & purification; Nitrilotriacetic Acid - chemistry; Protein Folding; Recombinant Proteins - biosynthesis; Recombinant Proteins - isolation & purification; Recombinant Proteins - therapeutic use; Tuberculosis - immunology; Tuberculosis - prevention & control; Tuberculosis Vaccines - biosynthesis; Tuberculosis Vaccines - isolation & purification; Tuberculosis Vaccines - therapeutic use; Vaccine; Vaccine; Mycobacterium tuberculosis; ESAT-6; Expression; MPT64
• ISSN: 1046-5928; 1096-0279
• DOI: 10.1016/j.pep.2007.11.016

The completion of Mycobacterium tuberculosis genome sequence has opened a new way for the identification and characterization of bacterial antigens, such as ESAT-6, CFP10, MPT64, and Ag85 complex, which are helpful for tuberculosis control. In this work, genes of ESAT-6 and MPT64 were fused and expressed in Escherichia coli in form of inclusion bodies with a histidine tag. The expressed fusion protein was purified by nitrilotriacetic acid (Ni–NTA) affinity chromatography under denaturing conditions, and the yield was 18 mg/L of culture. In mice, the purified ESAT-6–MPT64 fusion protein elicited stronger humoral response, greater splenic lymphocyte stimulated index, and higher levels of IFN-γ and IL-12 production than that of the single MPT64 inoculation group, and rendered modest protection on the experimental tuberculosis mouse models. In short, the ESAT-6–MPT64 fusion protein might be a potential candidate vaccine for tuberculosis.