Jeffrey G. Wright

The MerR heavy metal receptor mediates positive activation in a topologically novel transcription complex

Several physical and chemical signals from the extracellular environment are known to be transduced into changes in gene expression through multiple step pathways; however, mechanisms for triggering cellular responses to heavy metal stress have yet to be elucidated. We demonstrate here one such mechanism that employs a single heavy metal receptor protein, MerR, to directly activate transcription of the bacterial mercuric ion resistance operon. The mercuric ion-MerR complex and E. coli RNA polymerase holoenzyme synergistically bind to the metal responsive promoter in an unprecedented spatial relationship to form transcriptionally competent complexes. The activator binds adjacent to and overlaps with the polymerase molecule between the consensus -35 and -10 promoter regions. Our results support a model for transcriptional activation that includes both effector-induced protein-protein interactions and activator-induced alteration in DNA structure.

Biochemical and Spectroscopic Probes of Mercury(II) Coordination Environments in Proteins

Publisher Summary The high affinity of mercuric ion for thiolate ligands and the rapid ligand exchange rates of the resulting complexes make Hg(II) a relatively easy metal to bind to active sites of a variety of cysteine-containing enzymes. This feature has made mercuric ion a useful biochemical tool for selectively displacing one type of copper from a multicopper enzyme. It has become apparent that several spectroscopic features of Hg(II) complexes in their own right could be useful in distinguishing a variety of coordination environments. Although Hg(II) is a d 10 metal, it would be a mistake to consider it spectroscopically silent. As described here, the interaction of Hg(II) with biopolymers can be probed using extended X-ray absorption fine structure (EXAFS), UV-Vis, 199 Hg nuclear magnetic resonance (NMR), and circular dichroism (CD) spectroscopies. Several features in the spectra of structurally characterized model complexes have been correlated with the primary coordination number of the metal, aiding in the determination of coordination environments in proteins. This chapter discusses the advantages and limitations of using Hg(II) substitution in structure, function, and spectroscopic studies of proteins. Given these spectroscopic handles, determination of the coordination geometry, ligand identity, metal binding stoichiometry, dissociation rates, and binding constants are possible. Practical techniques, such as common methods for binding Hg(II) to proteins, are discussed. This chapter presents mercury chemistry, including structural and thermodynamic trends of biologically relevant mercury compounds.

XAS of the MerR metalloregulatory protein

The central component of a metal-responsive genetic switch, the MerR metalloregulatory protein, is one of the first examples of an mtracellular heavy metal receptor [l]. The merR gene product mediates the induction of the mercury resistance phenotype in bacteria [21, and resistant cells respond to subtoxic Hg(I1) levels (10 *6 -lo8 M) with transcriptional activation of the mer operon [31. Genetic evidence indicates that the merR gene product, MerR, exerts negative control of the structural genes in the absence of HgUI), and positive control in the presence of Hg(I1) 133.