1887

Abstract

Two-component systems play a central role in the adaptation of pathogenic bacteria to the environment prevailing within host tissues. The genes encoding the response regulator DevR (Rv3133c/DosR) and the cytoplasmic portion (DevS) of the histidine kinase DevS (Rv3132c/DosS), a putative two-component system of , were cloned and the protein products were overexpressed, purified and refolded as N-terminally His-tagged proteins from . DevS underwent autophosphorylation and participated in rapid phosphotransfer to DevR in a Mg-dependent manner. Chemical stability analysis and site-directed mutagenesis implicated the highly conserved residues His and Asp as the sites of phosphorylation in DevS and DevR, respectively. Mutations in Asp and Asp residues, postulated to form the acidic Mg-binding pocket, and the invariant Lys of DevR, abrogated phosphoryl transfer from DevS to DevR. DevR–DevS was thus established as a typical two-component regulatory system based on His-to-Asp phosphoryl transfer. Expression of the operon was induced at the RNA level in hypoxic cultures of H37Rv and was associated with an increase in the level of DevR protein. However, in a mutant strain expressing the N-terminal domain of DevR, induction was observed at the level of RNA expression but not at that of protein. DevS was translated independently of DevR and induction of transcripts was not associated with an increase in protein level in either wild-type or mutant strains, reflecting differential regulation of this locus during hypoxia.

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2004-04-01
2019-10-14
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