Replacement of the catalytic Zn(II) in horse liver alcohol dehydrogenase (HLADH) with copper produces a mononuclear Cu(II) chromophore with a ligand set consisting of two cysteine sulphurs, one histidine nitrogen plus one further atom. The fourth ligand to the metal ion and the conformation of the protein may be altered by addition of exogenous ligands and/or the cofactor NADH. Absorbance, CD, low-temperature magnetic CD (MCD) and EPR spectra are presented of copper-substituted HLADH samples in both ‘open’ and ‘closed’ conformations and in the presence and absence of the exogenous ligands pyrazole and DMSO. The EPR spectra indicate a strong, predominantly axial field about the copper(II) ion with high copper–thiol (cysteine) covalence. The optical and MCD spectra are interpreted in terms of four d-d transitions to low energy, also reflecting the axial ligand field, and four charge-transfer transitions to copper(II) between 30000 and 16000 cm-1 arising from the two cysteine sulphur atoms which give two pairs of oppositely signed MCD C-terms. These transitions are polarized mainly in the axial plane defined by Cys-46, Cys-174 and His-67. The binary complex formed with pyrazole displays quite different EPR and optical spectra which can be understood in terms of a rotation of the copper hole-orbital away from the axial plane thus decreasing sharply the copper–thiol covalence. The magneto-optical spectra in the presence and absence of DMSO are indistinguishable.
Magnetic and optical properties of copper-substituted alcohol dehydrogenase: a bisthiolate copper (II) complex
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Jacqueline A. FARRAR, Grazyna FORMICKA, Michael ZEPPEZAUER, Andrew J. THOMSON; Magnetic and optical properties of copper-substituted alcohol dehydrogenase: a bisthiolate copper (II) complex. Biochem J 15 July 1996; 317 (2): 447–456. doi: https://doi.org/10.1042/bj3170447
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