Characterization of Mycobacterium tuberculosis Rv3676 (CRP Mt ), a Cyclic AMP Receptor Protein-Like DNA Binding Protein

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Abstract
Little is known about cyclic AMP (cAMP) function in Mycobacterium tuberculosis , despite its ability to encode 15 adenylate cyclases and 10 cNMP-binding proteins. M. tuberculosis Rv3676, which we have designated CRP Mt , is predicted to be a cAMP-dependent transcription factor. In this study, we characterized CRP Mt 's interactions with DNA and cAMP, using experimental and computational approaches. We used Gibbs sampling to define a CRP Mt DNA motif that resembles the cAMP receptor protein (CRP) binding motif model for Escherichia coli . CRP Mt binding sites were identified in a total of 73 promoter regions regulating 114 genes in the M. tuberculosis genome, which are being explored as a regulon. Specific CRP Mt binding caused DNA bending, and the substitution of highly conserved nucleotides in the binding site resulted in a complete loss of binding to CRP Mt . cAMP enhanced CRP Mt 's ability to bind DNA and caused allosteric alterations in CRP Mt conformation. These results provide the first direct evidence for cAMP binding to a transcription factor in M. tuberculosis , suggesting a role for cAMP signal transduction in M. tuberculosis and implicating CRP Mt as a cAMP-responsive global regulator.