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

The environmental signals that affect gene regulation in Mycobacterium tube rculosis remain largely unknown despite their importance to tuberculosis pa thogenesis. Other work has shown that several promoters, including acr (als o known as hspX) (alpha -crystallin homolog), are upregulated in shallow st anding cultures compared with constantly shaking cultures. Each of these pr omoters is also induced to a similar extent within macrophages. The present study used two-dimensional get electrophoresis and mass spectrometry to fu rther characterize differences in mycobacterial protein expression during g rowth 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, a nd Acr were identified from each of four spots or gel bands that were speci fically increased in bacteria from standing cultures. An additional standin g-induced spot contained two comigrating proteins, GlcB and KatG. The great est induction was observed with Rv2623, a 32-kDa protein of unknown functio n that was strongly expressed under standing conditions and absent in shaki ng cultures. Analysis using PROBE, a multiple sequence alignment and databa se mining tool, classified M. tuberculosis Rv2623 as a member of a novel cl ass of ATP-binding proteins that may be involved in M. tuberculosis's respo nse to environmental signals. These studies demonstrate the power of combin ed proteomic and computational approaches and demonstrate that subtle diffe rences in bacterial culture conditions may have important implications for the study of gene expression in mycobacteria.