This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Salim, K. Articles by Huygen, K. Search for Related Content PubMed PubMed Citation Articles by Salim, K. Articles by Huygen, K. Agricola Articles by Salim, K. Articles by Huygen, K.

 Previous Article  |  Next Article 

Appl. Environ. Microbiol., 11 1997, 4392-4400, Vol 63, No. 11
Copyright © 1997, American Society for Microbiology

Heterologous expression of the Mycobacterium tuberculosis gene encoding antigen 85A in Corynebacterium glutamicum

K Salim, V Haedens, J Content, G Leblon and K Huygen
Laboratoire de Biologie Moleculaire des Corynebacteries, Institut de Genetique et de Microbiologie, URA 2225, Universite Paris-XI, Orsay, France.

By using appropriate Corynebacterium glutamicum-Escherichia coli shuttle plasmids, the gene encoding the fibronectin-binding protein 85A (85A) from Mycobacterium tuberculosis was expressed in C. glutamicum, also an actinomycete and nonsporulating gram-positive rod bacterium, which is widely used in industrial amino acid production. The 85A gene was weakly expressed in C. glutamicum under the control of the ptac promoter from E. coli, but it was produced efficiently under the control of the promoter of the cspB gene encoding PS2, one of the two major secreted proteins from C. glutamicum. The 85A protein was produced in various forms, with or without its own signal sequence and with or without the signal sequence and the NH2-terminal (18-amino- acid) mature sequence of PS2. Western blot analysis with monoclonal antibodies raised against the M. tuberculosis antigen 85 complex showed that recombinant 85A protein was present in the corynebacterial cell wall extract and also released in extracellular culture medium. NH2- terminal microsequencing of recombinant 85A secreted by C. glutamicum showed that signal peptide was effectively cleaved off at the predicted site. The recombinant 85A protein was biologically active in vitro, inducing significant secretion of Th1 T-cell cytokines, particularly interleukin-2 and gamma interferon, in spleen cell cultures from mice vaccinated with live Mycobacterium bovis BCG. Heterologous expression of mycobacterial antigens in C. glutamicum now offers a potent tool for further immunological characterization and large scale preparation of these recombinant proteins.

This article has been cited by other articles:

• Watanabe, K., Tsuchida, Y., Okibe, N., Teramoto, H., Suzuki, N., Inui, M., Yukawa, H. (2009). Scanning the Corynebacterium glutamicum R genome for high-efficiency secretion signal sequences. Microbiology 155: 741-750 [Abstract] [Full Text]  
• Kikuchi, Y., Itaya, H., Date, M., Matsui, K., Wu, L.-F. (2009). TatABC Overexpression Improves Corynebacterium glutamicum Tat-Dependent Protein Secretion. Appl. Environ. Microbiol. 75: 603-607 [Abstract] [Full Text]  
• Inui, M., Suda, M., Okino, S., Nonaka, H., Puskas, L. G., Vertes, A. A., Yukawa, H. (2007). Transcriptional profiling of Corynebacterium glutamicum metabolism during organic acid production under oxygen deprivation conditions. Microbiology 153: 2491-2504 [Abstract] [Full Text]  
• Vertes, A. A., Inui, M., Yukawa, H. (2005). Manipulating Corynebacteria, from Individual Genes to Chromosomes. Appl. Environ. Microbiol. 71: 7633-7642 [Full Text]  
• Gibson, K. J. C., Eggeling, L., Maughan, W. N., Krumbach, K., Gurcha, S. S., Nigou, J., Puzo, G., Sahm, H., Besra, G. S. (2003). Disruption of Cg-Ppm1, a Polyprenyl Monophosphomannose Synthase, and the Generation of Lipoglycan-less Mutants in Corynebacterium glutamicum. J. Biol. Chem. 278: 40842-40850 [Abstract] [Full Text]  
• Kikuchi, Y., Date, M., Yokoyama, K.-i., Umezawa, Y., Matsui, H. (2003). Secretion of Active-Form Streptoverticillium mobaraense Transglutaminase by Corynebacterium glutamicum: Processing of the Pro-Transglutaminase by a Cosecreted Subtilisin-Like Protease from Streptomyces albogriseolus. Appl. Environ. Microbiol. 69: 358-366 [Abstract] [Full Text]  
• Puech, V., Chami, M., Lemassu, A., Lanéelle, M.-A., Schiffler, B., Gounon, P., Bayan, N., Benz, R., Daffé, M. (2001). Structure of the cell envelope of corynebacteria: importance of the non-covalently bound lipids in the formation of the cell wall permeability barrier and fracture plane. Microbiology 147: 1365-1382 [Abstract] [Full Text]