Domain specificity in metal binding to metallothionein. A circular dichroism and magnetic circular dichroism study of cadmium and zinc binding at temperature extremes.
Rabbit liver Zn metallothionein-(MT) will bind cadmium readily between -26 degrees C and 70 degrees C. The binding reaction was monitored by recording the circular dichroism and magnetic circular dichroism spectra, in the region of the RS(-)----Cd2+ charge transfer transition at 250 nm, at intervals as aliquots of cadmium were added. For all temperatures, these data can be analyzed in terms of a distributed mechanism for cadmium binding when Zn-MT is used, and a domain-specific mechanism when apo-MT is used. The CD spectrum measured at -26 degrees C for Cd,Zn-MT, which was made by adding excess cadmium directly to Zn7-MT at -26 degrees C, is not the same as the CD spectrum of Cd-MT prepared at room temperature from the same Zn7-MT. Measurements of the stoichiometry of the cadmium and zinc bound to MT in the presence of excess cadmium at different temperatures indicates that below 5 degrees C at least one zinc atom remains bound to the protein. The mixed metal metallothionein, Cd/Zn-MT, that always forms below 5 degrees C, is characterized by a single maximum near 250 nm in the CD spectrum, rather than the derivative-shaped CD envelope that is diagnostic of the (Cd4-S11)alpha cluster, which indicates that the zinc occupies a site in the alpha domain. Rearrangement of the bound metals to the domain-specific distribution takes place if Cd,Zn-MT, prepared at subzero temperatures, is warmed above 30 degrees C.
Stillman
M J
MJ
Department of Chemistry, University of Western Ontario, London, Canada.
Zelazowski
A J
AJ
D000818:Animals; D001665:Binding Sites; D002104:Cadmium / Q000378:metabolism*; D002614:Chelating Agents; D002942:Circular Dichroism; D008099:Liver / Q000378:metabolism; D008280:Magnetics; D008668:Metallothionein / Q000378:metabolism*; D008670:Metals / Q000378:metabolism*; D011817:Rabbits; D012117:Resins, Synthetic; D013696:Temperature; D015032:Zinc / Q000378:metabolism*
D002614:Chelating Agents; D008670:Metals; D012117:Resins, Synthetic; D002104:Cadmium; C024997:Chelex 100; D008668:Metallothionein; D015032:Zinc