Accession Number : AD1011470

Title :   Identification and Characterization of Functional Domains of the Diphtheria Toxin Repressor (DtxR)

Descriptive Note : Technical Report

Corporate Author : Uniformed Services University Of The Health Sciences Bethesda United States

Personal Author(s) : Wang,Zhao-Xi

Full Text :

Report Date : 01 Jan 1995

Pagination or Media Count : 178

Abstract : Corynebacterium diphtheriae is the causative agent of diphtheria. Diphtheria toxin, a potent bacterial exotoxin encoded by the tox gene of certain temperate corynebacteriophages, is the primary virulence factor of C. diphtheriae. In C. diphtheriae, the production of diphtheria toxin and the iron uptake system are negatively controlled by the diphtheria toxin repressor (DtxR), an iron-dependent metalloregulatory protein. The regulatory function of DtxR can be activated by various divalent cations, including Fe2+, Cd2+, Co2+, Mn2+, Ni2+, and Zn2+.In this investigation, we focused on understanding the structural basis of metalloregulation of DtxR. First, we identified and characterized 20 distinct mutations in dtxR induced by random mutagenesis with bisulfite treatment, eighteen of which caused single amino acid substitutions in DtxR and two of which were chain-terminating mutations. Six of the amino acid replacements were clustered between residues 39 and 52 in a predicted helix-turn-helix motif that exhibits homology with several other repressors and is identified as the putative DNA-binding domain of DtxR. Three substitutions occurred within a predicted alpha-helical region with the sequence His98-X3-Cys102-X3-His1Q6 that resembles metal-binding motifs in several other proteins and is identified as the putative metal-binding site of DtxR. Second, we targeted the probable metal-coordinating histidine and cysteine residues within the metal-binding sequence of DtxR for oligonucleotide-directed site-specific mutagenesis and isolated six additional DtxR variants.

Descriptors :   Corynebacterium diptheriae , exotoxins , iron , proteins , mutations , amino acids , Histidine , Cysteine , Cytotoxin , toxicity , deoxyribonucleic acids , assaying , purification , chromatography

Distribution Statement : APPROVED FOR PUBLIC RELEASE