An inducible expression system for high-level expression of recombinant proteins in slow growing mycobacteria.
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| Abstract |
A novel protein expression vector utilising the inducible hspX promoter of Mycobacterium tuberculosis was constructed and evaluated in this study. High-level induction of three mycobacterial antigens, comprising up to 9% of bacterial sonicate, was demonstrated in recombinant Mycobacterium bovis BCG when grown under low-oxygen tension, which serves to enhance hspX promoter activity. Recombinant proteins were efficiently purified from bacterial lysates in a soluble form by virtue of a C-terminal 6-histidine tag. Purification of the immunodominant M. tuberculosis Ag85B antigen using this system resulted in a recombinant protein that stimulated significant IFN-γ release from Ag85B-reactive T cells generated after vaccination of mice with an Ag85B-expressing vaccine. Further, the M. tuberculosis L-alanine dehydrogenase (Ald) protein purified from recombinant BCG displayed strong enzymatic activity in recombinant form. This study demonstrated that high levels of native-like recombinant mycobacterial proteins can be produced in mycobacterial hosts, and this may aid the analysis of mycobacterial protein function and the development of new treatments.
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| Authors | Lisa Leotta, Joanne M Spratt, Carlyn U Kong, James A Triccas |
| Journal | Plasmid (Plasmid) Vol. 81 Pg. 27-31 (Sep 2015) ISSN: 1095-9890 [Electronic] United States |
| PMID | 26021569 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't) |
| Copyright | Copyright © 2015 Elsevier B.V. All rights reserved. |
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