Cadmium-109 as a probe of the metal binding sites in horse liver alcohol dehydrogenase

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Abstract
The noncatalytic and catalytic Zn atoms of horse liver alcohol dehydrogenase, [(LADH)Zn2Zn2] or LADH, were replaced differentially with 109Cd by equilibrium dialysis, resulting in 2 new enzymatically active species, [(LADH)109Cd2Zn2] and [(LADH)109Cd2109Cd2]. The UV difference spectra of the Cd enzymes vs. native [(LADH)Zn2Zn2] reveal maxima at 240 nm with molar absorptivities, .DELTA..epsilon.240, of 1.6 .times. 104 M-1 cm-1 per noncatalytic 109Cd atom and 0.9 .times. 104 M-1 cm-1 per catalytic 109Cd atom, consistent with coordination of the metals by 4 and 2 thiolate ligands, respectively, strikingly similar to the 250-nm charge-transfer absorbance in metallothionein. Carboxymethylation of the Cys-46 ligand to the catalytic metal in LADH presumably lowers the overall stability constant of the coordination complex and results in loss of catalytic 109Cd or catalytic Co but not catalytic Zn from the enzyme.