Analysis of genes that encode DtxR-like transcriptional regulators in pathogenic and saprophytic corynebacterial species

Metal-dependent transcriptional regulators of the diphtheria toxin repressor (DtxR) family have been identified in a wide variety of bacterial genera, where they control gene expression in response to one of two metal ions, Fe2+ or Mn2+. DtxR of Corynebacterium diphtheriae is the best characterized of these important metal-dependent regulators. The genus Corynebacterium includes many phenotypically diverse species, and the prevalence of DtxR-like regulators within the genus is unknown. We assayed chromosomal DNA from 42 different corynebacterial isolates, representing 33 different species, for the presence of a highly conserved region of the dtxR gene that encodes the DNA-binding helix-turn-helix motif and metal-binding site I within domains 1 and 2 of DtxR. The chromosome of all of the isolates contained this conserved region of dtrR, and DNA sequencing revealed a high level of nucleotide sequence conservation within this region in all of the corynebacterial species (ranging from 62 to 100% identity and averaging 70% identity with the dtrR prototype). The level of identity was even greater for the predicted protein sequences encoded by the dtxR-like genes, ranging from 81 to 100% identity and averaging 91% identity with DtxR. Using a DtxR-specific antiserum we confirmed the presence of a DtxR-like protein in extracts of most of the corynebacterial isolates and determined the precise amount of DtxR per cell in C diphtheriae. The high level of identity at both DNA and protein levels suggests that all of the isolates tested encode a functional DtxR-like Fe2+-activated regulatory protein that can bind homologs of the DtxR operator and regulate gene expression in response to iron.