MerR protein

The detection of metal-binding proteins, especially of Cd- and Hg-binding metallothioneins or of the merR protein, induced numerous studies of model compounds of Cd and Hg with more or less simple sulfur- and selenium-containing ligands. 

Since Hg(II) has a closed-shell electronic structure, it is often considered spectroscopically silent in optical and EPR spectroscopy. Recent advances in a variety of spectroscopic techniques have made this description of Hg(II) complexes obsolete. Relativistic effects tend to lower energy of the Hg 6s orbital (156), and consequently the LMCT spectra of Hg(II) tetrahalides and three- or four-coordinate alkyl thiolate complexes are distinctly different than those obtained for two-coordinate complexes. As described below, ultraviolet spectroscopy has provided details about the Hg(II) coordination in the MerR protein (202, 210). 

Figure 9. Lowest energy LMCT bands of several proteins plotted as a function of ligand optical electronegativity. For comparison, the lowest energy bands of tetrahedral halide Hg(II) complexes arc shown, along with a line obtained from the halide data. As bis Hg(II)-thiolate complexes have significantly higher LMCT energies, these data suggest that Hg(II) in the MerR protein is in a higher number coordination environment.

Figure 14. Sequences of the five reported MerR proteins. The Tn21 and TnSOl MerR proteins arc from narrow-spectrum resistance systems to mercuric ion, while the pDU1358,...

RC607, and pI258 MerR proteins come from broad-spectrum mercuric ion and organom-ercurial resistance systems (82, 176, 208). Bold residues indicate the residues that are strictly conserved among the five proteins. Bold, double underlined residues highlight the three cysteinyl residues believed to be involved in Hg(II) recognition and transcriptional activation. 

The overexpression and purification of the dimeric, 144 amino acid TnSOl MerR protein in large quantities has made detailed chemical studies possible (143, 145, 171). A variety of studies including gel filtration, equilibrium dialysis, nonequilibrium dialysis, and spectrophotometric titration have indicated that the MerR proteins isolated to date bind a single mercuric ion per protein dimer (82, 145, 171, 202). All of these studies have been performed in the presence of 10-10,(XX)-fold excess buffer thiol competitors, indicating that Hg(II) binds tightly to the protein. 

Figure 16. General models for Hg(II) binding to the MerR protein. Mutagenesis studies led to the proposal of model 1 having linear bis coordination with ancillary ligation (173). Studies by electronic spectroscopy and Hg-EXAFS, indicate that Hg(II) is bound by three primary thiolate ligands, consistent with models 2 and 3, above (202, 209). Mutant complementation studies with Bacillus MerR support the model for Hg(II) bound by three primary bonds (83).

A reaction scheme, elaborated from Eq. 10, for initiation of transcription in the presence and absence of mercuric ion is presented in Fig. 17. Comparison of the kinetic constants for each state has shown that Hg(II) binding to the MerR protein results in an increase in the isomerization rate constant kf) of RNA polymerase of almost two orders of magnitude, whereas the binding ( b) of RNAP to the promoter is not greatly affected (158). 

Hg(II) binding induces a conformational change in the MerR protein, as demonstrated by a three fold decrease in the affinity of MerR for DNA in the presence of Hg(II). 

Dimer complementation studies of site-directed cysteinyl mutants for the Bacillus RC607 MerR protein, as discussed in Sections VIIB and C, have recently been reported (83a). 

A resolution, respectively) have also been determined. The metal coordination environment of these proteins is likely essential for determining metal specificity of MerR proteins. 

The expression of proteins involved in Hg(II) detoxification is regulated by the MerR protein. The MerR protein is always bound as a dimer adjacent to the RNA polymerase binding site of the mer gene. In the absence of Hg, MerR holds the DNA in a conformation so that the RNA polymerase binding is blocked and transcription cannot occur. When the mercury binds to MerR, it changes the conformation of the MerR protein-DNA complex and allows RNA polymerase to bind and transcribe the mer operon, creating mRNA for the series of enzymes that carry out mercury resistance. 

In the absence of Hg(II), the transcription and synthesis of these Hg detoxification enzymes is inhibited by a regulatory protein, called merR, that binds to a specific location in the mer operon, the section of DNA coding for mercury resistance. When Hg(il) is present, it binds to three Cys residues of the merR protein. This causes a conformational change in both the protein and in the DNA to which it is bound that leads to transcription of the lyase and reductase. In this way, these proteins are only produced when required. ... 

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