Abstract
The Staphylococcus aureus plasmid pI258 cadCA operon encodes a P-type ATPase, CadA, that confers resistance to Cd(II)/Pb(II)/Zn(II). Expression is regulated by CadC, a homodimeric repressor that dissociates from the cad operator/promoter upon binding of Cd(II), Pb(II), or Zn(II). CadC is a member of the ArsR/SmtB family of metalloregulatory proteins. The crystal structure of CadC shows two types of metal binding sites, termed Site 1 and Site 2, and the homodimer has two of each. Site 1 is the physiological inducer binding site. The two Site 2 metal binding sites are formed at the dimerization interface. Site 2 is not regulatory in CadC but is regulatory in the homologue SmtB. Here the role of each site was investigated by mutagenesis. Both sites bind either Cd(II) or Zn(II). However, Site 1 has higher affinity for Cd(II) over Zn(II), and Site 2 prefers Zn(II) over Cd(II). Site 2 is not required for either derepression or dimerization. The crystal structure of the wild type with bound Zn(II) and of a mutant lacking Site 2 was compared with the SmtB structure with and without bound Zn(II). We propose that an arginine residue allows for Zn(II) regulation in SmtB and, conversely, a glycine results in a lack of regulation by Zn(II) in CadC. We propose that a glycine residue was ancestral whether the repressor binds Zn(II) at a Site 2 like CadC or has no Site 2 like the paralogous ArsR and implies that acquisition of regulatory ability in SmtB was a more recent evolutionary event.
Highlights
Encodes at least two gene products; that is, a transcriptional repressor and a resistance protein such as an efflux pump, intracellular chelator, or modifying enzyme
The cadCA operon of Staphylococcus aureus plasmid pI258 confers resistance to Cd(II), Pb(II), and Zn(II) and encodes two proteins: CadC, a metal-responsive transcriptional repressor that responds to Cd(II), Pb(II), and Zn(I) [3,4,5] and CadA, a P-type ATPase that extrudes the soft metals out of the cell [6]
The crystal structure of apo-CadC revealed a second type of binding site, termed Site 2, at the C-terminal interface between the dimers is composed of four residues, Asp101 and His103 from one monomer and His114 and Glu117 from the other monomer [9]
Summary
Encodes at least two gene products; that is, a transcriptional repressor (metal sensor protein) and a resistance protein such as an efflux pump, intracellular chelator, or modifying enzyme. CadC has two different types of metal binding sites (four binding sites per dimer) [7, 8]. The crystal structure of apo-CadC revealed a second type of binding site, termed Site 2, at the C-terminal interface between the dimers is composed of four residues, Asp101 and His103 from one monomer and His114 and Glu117 from the other monomer [9]. In this study the Site 1 and Site 2 metal binding sites were eliminated by mutagenesis. The stoichiometry, relative affinities, and coordination geometry of both sites of were determined Both types of sites bound both Cd(II) and Zn(II), but Site 1 had a preference for Cd(II) over Zn(II), and the reverse was found for Site 2. Mutagenesis of the metal binding residues in Site 2 eliminated binding of Zn(II) at the dimer interface, there was only a minimal effect on dimerization. The results suggest that a single residue at position 87 may be responsible for the difference between the non-regulatory Zn(II) site in CadC and the Zn(II)-responsive site in SmtB
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
