Cd(II)-responsive and constitutive mutants implicate a novel domain in MerR

Expression of the Tn21 mercury resistance (mer) operon is controlled by a m etal-sensing repressor-activator, MerR. When present, MerR always binds to the same position on the DNA (the operator merO), repressing transcription of the structural genes merTPCAD in the absence of Hg(II) and inducing thei r transcription in the presence of Hg(II). Although it has two potential bi nding sites, the purified MerR homodimer binds only one Hg(II) ion, employi ng Cys82 from one monomer and Cyr117 and Cys126 from the other. When MerR b inds Hg(PI), it changes allosterically and also distorts the merO DNA to fa cilitate transcriptional initiation by sigma(70) RNA polymerase. Wild-type MerR is highly specific for Hg(II) and is 100- and 1,000-fold less responsi ve to the chemically related group 12 metals, Cd(II) and Zn(II), respective ly. We sought merR mutants that respond to Cd(II) and obtained 11 Cd(II)-re sponsive and 5 constitutive mutants. The Cd(II)-responsive mutants, most of which had only single-residue replacements, were also repression deficient and still Hg(II) responsive but, like the wild type, were completely unres ponsive to Zn(II). None of the Cd(II)-responsive mutations occurred in the DNA binding domain or replaced any of the key Cys residues. Five Cd(II)-res ponsive single mutations lie in the antiparallel coiled-coil domain between Cys82 and Cys117 which constitutes the dimer interface. These mutations id entify 10 new positions whose alteration significantly affect MerR's metal responsiveness or its repressor function. They give rise to specific predic tions for how MerR distinguishes group 12 metals, and they refine our model of the novel domain structure of MerR. Secondary-structure predictions sug gest that certain elements of this model also apply to other MerR family re gulators.