metallidurans (PbrR: [15, 56]) or using FRET (PbrR691, [13]) without any transcriptional response to Zn or Cd, whereas related MerR family regulators that have been tested respond to a greater or lesser extent to Zn(II), Cd(II) and Pb(II) [10, 23, 57], as do SmtB/ArsR family repressors [47, 54]. However, transcriptomics experiments indicate that the pbr structural genes are also induced in the presence of other metals, arguing
that expression of the pbr operon and other metal resistance operons in C. metallidurans is influenced by other factors [7, 12]. Our experiments show that the mechanism of transcriptional activation by PbrR appears selleck chemical to be essentially identical to that of MerR family regulators that have been characterized. PbrR contains three cysteine LXH254 in vivo residues that are necessary for Pb(II)-induced transcription from the pbrA promoter. C14 is in the helix-turn-helix DNA binding domain, and may be essential for the regulator/DNA interaction. C79 is essential in all divalent metal ion responsive MerR regulators tested so far, whilst C134 is not found in other characterized MerR
regulators. Our data show that PbrR transcription is activated by Pb(II) using different amino acids to other divalent metal ion-activated MerR regulators, but further work is required to determine G418 ic50 whether Pb(II) coordinates other residues in PbrR. Acknowledgements We gratefully acknowledge the contribution of Niels van der Lelie and Brigitte Borremans to the start of this project and to Max Mergeay for advice and training to DJJ. We thank Chris Kershaw for critical reading of the manuscript. This work was supported by the Biotechnology and Biological Sciences Research Council (research grant B10333
and a studentship to DJJ). The Birmingham Functional Genomics laboratory was supported by a Joint Infrastructure Fund grant JIF13209 and bioinformatics facilities were provided through MRC Infrastructure Award G.4600017. References 1. Mire CE, Tourjee JA, O’Brien WF, Ramanujachary KV, Hecht GB: Lead precipitation by Vibrio harveyi: evidence for novel quorum-sensing interactions. Appl Environ Microbiol 2004, 70:855–864.PubMedCrossRef 2. Rensing C, Sun Y, Mitra PDK4 B, Rosen BP: Pb(II)-translocating P-type ATPases. J Biol Chem 1998, 49:32614–32617.CrossRef 3. Sharma R, Rensing C, Rosen BP, Mitra B: The ATP hydrolytic activity of purified ZntA, a Pb(II)/Cd(II)/Zn(II)-translocating ATPase from Escherichia coli. J Biol Chem 2000, 275:3873–3878.PubMedCrossRef 4. Borremans B, Hobman JL, Provoost A, Corbisier P, Brown NL, van der Lelie D: Cloning and functional analysis of the pbr lead resistance determinant of Ralstonia metallidurans CH34. J Bacteriol 2001, 183:5651–5658.PubMedCrossRef 5.