Specificity of the Metalloregulator CueR for Monovalent Metal Ions: Possible Functional Role of a Coordinated Thiol?

Abstract

Metal-ion-responsive transcriptional regulators within the MerR family effectively discriminate between mono- and divalent metal ions. Herein we address the origin of the specificity of the CueR protein for monovalent metal ions. Several spectroscopic techniques were employed to study Ag(I) , Zn(II) , and Hg(II) binding to model systems encompassing the metal-ion-binding loop of CueR from E. coli and V. cholerae. In the presence of Ag(I) , a conserved cysteine residue displays a pKa value for deprotonation of the thiol that is close to the physiological pH value. This property is only observed with the monovalent metal ion. Quantum chemically optimized structures of the CueR metal site with Cys 112 protonated demonstrate that the conserved Ser 77 backbone carbonyl oxygen atom from the other monomer of the homodimer is "pulled" towards the metal site. A common allosteric mechanism of the metalloregulatory members of the MerR family is proposed. For CueR, the mechanism relies on the protonation of Cys 112.

Metallochaperones and metalloregulation in bacteria

“…The functional specificity of a single sensing site is achieved by precise remodelling of coordination number, ligand type and geometry in MerRs [103,107,108]. MerRs bind inducing metals ions via coordination by a minimum of two conserved cysteine residues present in a C-terminal metal-binding loop, and Cu I , Ag I and Au I all adopt this bis -thiolato coordination structure, e.g.…”

Metalloproteinek fémkötőhelyein alapuló oligopeptidek, mint potenciális toxikus fémion érzékelők

“…28 The major result of our studies on the various metal ionoligopeptide systems is a finding that the model ligands possess fundamentally distinct binding features for the monovalent ion Ag + and the divalent ions Zn 2+ , Cd 2+ and Hg 2+ . [29][30][31][32] The binding of Ag + to the ligands under acidic conditions (even at pH = 2) was proved by UV-, CD-and 1 H-NMR experiments, as well as pH-potentiometric titrations ( Figure 6). 31 Spectroscopic data suggest the coordination of at least one of the Cys-sidechain donors as a thiolate to Ag + in the Ag + -PP 1:1 system (PP = Ac-SCPGDQGSDCPI-NH 2 ).…”

“…[29][30][31][32] The binding of Ag + to the ligands under acidic conditions (even at pH = 2) was proved by UV-, CD-and 1 H-NMR experiments, as well as pH-potentiometric titrations ( Figure 6). 31 Spectroscopic data suggest the coordination of at least one of the Cys-sidechain donors as a thiolate to Ag + in the Ag + -PP 1:1 system (PP = Ac-SCPGDQGSDCPI-NH 2 ). However, a deprotonation process in the pH-range of pH~5.5 -8 can be clearly observed both on the pH-and UV-titration curves (Figure 6.)…”

“…Indeed, a {Cys-S -,Cys-SH} type coordination environment around the monovalent metal ions is also possible in the metal binding site of CueR. 31 These results are an important step towards the deeper understanding of the mechanism of operation of the protein. At the same time, our findings also provide an opportunity for utilizing the complex recognition mechanism of the protein in the development of genetically modified bacteria that produce a fluorescence signal as a response to a specific metal ion and thus behave as a metal-responsive biosensor.…”

“…37 A {Cys-S -,Cys-SH} típusú kötésmód CueR fehérje fémkötõ centrumában is racionális lehetõség, aminek jelentõs szerepe lehet a molekula mûködési mechanizmusában. 31 …”