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Infection and Immunity, September 1998, p. 4208-4214, Vol. 66, No. 9
0019-9567/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Molecular Cloning, Expression, and Immunogenicity of MTB12, a Novel Low-Molecular-Weight Antigen Secreted by Mycobacterium tuberculosis

John R. Webb,1,dagger Thomas S. Vedvick,2 Mark R. Alderson,2 Jeffrey A. Guderian,2 Shyian S. Jen,2 Pamela J. Ovendale,2 Stephen M. Johnson,1,Dagger Steven G. Reed,1,2,3 and Yasir A. W. Skeiky2,*

Infectious Disease Research Institute1 and Corixa Corporation,2 Seattle, Washington 98104, and Department of Pathobiology, University of Washington, Seattle, Washington 981953

Received 4 March 1998/Returned for modification 20 April 1998/Accepted 10 June 1998

Proteins secreted into the culture medium by Mycobacterium tuberculosis are thought to play an important role in the development of protective immune responses. In this report, we describe the molecular cloning of a novel, low-molecular-weight antigen (MTB12) secreted by M. tuberculosis. Sequence analysis of the MTB12 gene indicates that the protein is initially synthesized as a 16.6-kDa precursor protein containing a 48-amino-acid hydrophobic leader sequence. The mature, fully processed form of MTB12 protein found in culture filtrates has a molecular mass of 12.5 kDa. MTB12 protein constitutes a major component of the M. tuberculosis culture supernatant and appears to be at least as abundant as several other well-characterized culture filtrate proteins, including members of the 85B complex. MTB12 is encoded by a single-copy gene which is present in both virulent and avirulent strains of the M. tuberculosis complex, the BCG strain of M. bovis, and M. leprae. Recombinant MTB12 containing an N-terminal six-histidine tag was expressed in Escherichia coli and purified by affinity chromatography. Recombinant MTB12 protein elicited in vitro proliferative responses from the peripheral blood mononuclear cells of a number of purified protein derivative-positive (PPD+) human donors but not from PPD- donors.

* Corresponding author. Mailing address: Corixa Corporation, 1124 Columbia St., Suite 200, Seattle, WA 98104. Phone: (206) 754-5772. Fax: (206) 754-5715. E-mail: skeiky{at}corixa.com.

dagger Present address: Department of Microbiology and Immunology, University of Ottawa, Ottawa, Canada K1H 8M5.

Dagger Present address: P.O. Box 1944, Chinle, AZ 86503.

Infection and Immunity, September 1998, p. 4208-4214, Vol. 66, No. 9
0019-9567/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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