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Infection and Immunity, September 2001, p. 5777-5785, Vol. 69, No. 9
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.9.5777-5785.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Identification and Characterization of Mycobacterial Proteins Differentially Expressed under Standing and Shaking Culture Conditions, Including Rv2623 from a Novel Class of Putative ATP-Binding Proteins

Matthew A. Florczyk,1,2 Lee Ann McCue,2 Robert F. Stack,2 Charles R. Hauer,1,2 and Kathleen A. McDonough1,2,*

Department of Biomedical Sciences, University at Albany, Albany, New York 12222,1 and Wadsworth Center, New York State Department of Health, Albany, New York 12202-20022

Received 15 February 2001/Returned for modification 2 April 2001/Accepted 4 June 2001

The environmental signals that affect gene regulation in Mycobacterium tuberculosis remain largely unknown despite their importance to tuberculosis pathogenesis. Other work has shown that several promoters, including acr (also known as hspX) (alpha -crystallin homolog), are upregulated in shallow standing cultures compared with constantly shaking cultures. Each of these promoters is also induced to a similar extent within macrophages. The present study used two-dimensional gel electrophoresis and mass spectrometry to further characterize differences in mycobacterial protein expression during growth under standing and shaking culture conditions. Metabolic labeling of M. bovis BCG showed that at least 45 proteins were differentially expressed under standing and shaking culture conditions. Rv2623, CysA2-CysA3, Gap, and Acr were identified from each of four spots or gel bands that were specifically increased in bacteria from standing cultures. An additional standing-induced spot contained two comigrating proteins, GlcB and KatG. The greatest induction was observed with Rv2623, a 32-kDa protein of unknown function that was strongly expressed under standing conditions and absent in shaking cultures. Analysis using PROBE, a multiple sequence alignment and database mining tool, classified M. tuberculosis Rv2623 as a member of a novel class of ATP-binding proteins that may be involved in M. tuberculosis's response to environmental signals. These studies demonstrate the power of combined proteomic and computational approaches and demonstrate that subtle differences in bacterial culture conditions may have important implications for the study of gene expression in mycobacteria.

* Corresponding author. Mailing address: Wadsworth Center, New York State Department of Health, P.O. Box 22002, Albany, NY 12202-2002. Phone: (518) 486-4253. Fax: (518) 474-3181. E-mail: Kathleen.McDonough{at}wadsworth.org.

Infection and Immunity, September 2001, p. 5777-5785, Vol. 69, No. 9
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.9.5777-5785.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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