title: Mycobacterium tuberculosis antigen 85B and ESAT-6 expressed as a recombinant fusion protein in Mycobacterium smegmatis elicits cell-mediated immune response in a murine vaccination model.
abstract: In this study, we investigated the potential molecular and immunological differences of a recombinant fusion protein (Hybrid-1), comprising of the immunodominant antigens Ag85B and ESAT-6 from Mycobacterium tuberculosis, derived from two different expression systems, namely Mycobacterium smegmatis and Escherichia coli. The fusion protein was successfully expressed and purified from both bacterial hosts and analyzed for any host-dependent post-translational modifications that might affect the immunogenicity of the protein. We investigated the immunogenicity of Hybrid-1 expressed in the two host species in a murine vaccination model, together with a reference standard Hybrid-1 (expressed in E. coli) from the Statens Serum Institut. No evidence of any post-translation modification was found in the M. smegmatis-derived Hybrid-1 fusion protein, nor were there any significant differences in the T-cell responses obtained to the three antigens analyzed. In conclusion, the Hybrid-1 fusion protein was successfully expressed in a homologous expression system using M. smegmatis and this system is worth considering as a primary source for vaccination trials, as it provided protein of excellent yield, stability and free from lipopolysaccharide.
authors: Centre for Infection, Immunity and Disease Mechanisms, Biosciences, School of Health Sciences and Social Care, Brunel University, London UB8 3PH, United Kingdom. anthony.tsolaki@brunel.ac.uk
authors: Anthony G
authors: AG
authors: Tsolaki
authors: Judit
authors: J
authors: Nagy
authors: Sergio
authors: S
authors: Leiva
authors: Uday
authors: U
authors: Kishore
authors: Ida
authors: I
authors: Rosenkrands
authors: Brian D
authors: BD
authors: Robertson
mesh_terms: D000217:Acyltransferases / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D000818:Animals; D000907:Antibodies, Bacterial / Q000235:genetics / Q000276:immunology; D000917:Antibody Formation / Q000235:genetics / Q000276:immunology*; D000942:Antigens, Bacterial / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D003001:Cloning, Molecular / Q000379:methods; D004926:Escherichia coli / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015870:Gene Expression / Q000276:immunology; D007111:Immunity, Cellular / Q000235:genetics / Q000276:immunology; D051379:Mice; D020102:Mycobacterium smegmatis / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism; D032581:Tuberculosis Vaccines / Q000235:genetics* / Q000276:immunology
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C093276:ESAT-6 protein, Mycobacterium tuberculosis; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D032581:Tuberculosis Vaccines; D000217:Acyltransferases; C121433:antigen 85B, Mycobacterium tuberculosis