Roles of the four cysteine residues in the function of the integral inner membrane Hg2+-binding protein, MerC.
The roles of the four cysteine residues of the integral inner membrane Hg2+-binding protein, MerC, have been examined using site-directed mutagenesis. Residues Cys-22 and Cys-25 have previously been predicted to lie within the membrane. Substitution of each of these residues in turn with alanine resulted in complete abolition of specific Hg2+ uptake by vesicles. In contrast, substitution by alanine of the other two cysteine residues, Cys-127 and Cys-132, predicted to lie with within a C-terminal cytoplasmic tail, did not significantly affect Hg2+ uptake. Since previous results indicated that native MerC tends to form intermolecular disulfide-bonded dimers, the effects of these substitutions on dimer formation were also examined. Only the Cys-127 and Cys-132 variants spontaneously formed significant amounts of disulfide-bonded dimer. Further experiments using copper-1,10-phenanthroline indicated that each variant with an unpaired cysteine residue was more susceptible to dimer formation than native MerC.
Sahlman
L
L
Department of Biochemistry, University of Umeå, Sweden. Lena@chem.umu.se
Hägglöf
E M
EM
Powlowski
J
J
D000818:Animals; D001426:Bacterial Proteins; D002352:Carrier Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D027682:Cation Transport Proteins; D003545:Cysteine / Q000737:chemistry* / Q000502:physiology; D004926:Escherichia coli; D008565:Membrane Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D008628:Mercury / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding / Q000235:genetics; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism
D001426:Bacterial Proteins; D002352:Carrier Proteins; D027682:Cation Transport Proteins; D008565:Membrane Proteins; D011994:Recombinant Proteins; C072715:merC protein, Acidithiobacillus ferrooxidans; D008628:Mercury; D003545:Cysteine