Evidence for noncooperative metal binding to the alpha domain of human metallothionein.
In the present study, we investigated the metal-binding reactivity of the isolated alpha domain of human metallothionein isoform 1a, with specific emphasis on resolving the debate concerning the cooperative nature of the metal-binding mechanism. The metallation reaction of the metal-free alpha domain with Cd2+ was unequivocally shown to proceed by a noncooperative mechanism at physiologic pH by CD and UV absorption spectroscopy and ESI MS. The data clearly show the presence of intermediate partially metallated metallothionein species under limiting Cd2+ conditions. Titration with four molar equivalents of Cd2+ was required for the formation of the Cd4alpha species in 100% abundance. The implications of a noncooperative metal-binding mechanism are that the partially metallated and metal-free species are stable intermediates, and thus may have a potential role in the currently undefined function of metallothionein.
Rigby Duncan
Kelly E
KE
Department of Chemistry, The University of Western Ontario, London, ON, Canada.
Stillman
Martin J
MJ
D000595:Amino Acid Sequence; D002104:Cadmium / Q000737:chemistry / Q000378:metabolism; D002413:Cations, Divalent; D006801:Humans; D008668:Metallothionein / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D019216:Metals, Heavy / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D020033:Protein Isoforms / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000138:chemical synthesis / Q000378:metabolism
D002413:Cations, Divalent; D019216:Metals, Heavy; D010446:Peptide Fragments; D020033:Protein Isoforms; D011994:Recombinant Proteins; C407523:metallothionein isoform 1; D002104:Cadmium; D008668:Metallothionein