Bovine copper-zinc superoxide dismutase active site

Kamnakaran, C., Zhang, H., Crow, J.P., Anthohne, W.E., Kalyanaraman, B., 2004. Direct probing of copper active site and free radical formed during bicarbonate-dependent peroxidase activity of bovine and human copper, zinc-superoxide dismutases. Low-temperature electron paramagnetic resonance and electron nuclear double resonance studies. J. Biol. Chem. 279, 32534—32540. 

The structure and enzyme kinetics of bovine erythrocyte superoxide dismutase are reviewed. The protein has a novel imidazolate-bridged copper(II)-zinc(II) catalytic center in each of two identical subunits. Since a C /Cu1 redox couple is responsible for the dismutase activity of the enzyme, the role of zinc is of interest. Both 220-MHz NMR measurements of the exchangeable histidine protons and chemical modifications using diethylpyrocarbonate demonstrate that zinc alone can fold the protein chain in the region of the active site into a conformation resembling that of the native enzyme. Other possible roles for zinc are discussed. Synthetic, magnetic, and structural studies of soluble, imidazolate-bridged copper complexes of relevance to the 4 Cu(II) form of the enzyme have been made. 

Superoxide dismutase catalyzes the disproportionation of superoxide as 2O2 -i- 2H — O2 -I- H2O2. The Cu,Zn-superoxide dismutases are widely distributed in both plant and animal kingdoms and are found in eukaryotic cytosols. The vast majority of studies on superoxide dismutases have been made on the enzyme from bovine erythrocytes this enzyme contains 151 amino acid residues and two copper and two zinc atoms per molecule of —32,000 Da. The crystal structure of this protein has been reported (97), and the active site was shown to consist of a bimetallic (Cu,Zn) assembly (see Fig. 6). A type-2 copper center is... 

It has been concluded from a study of the optical and e.p.r. spectra of Co —Cu bovine superoxide dismutase, in which zinc has been replaced by cobalt, that the cobalt site reactivity should be described in terms of reaction of the Co-imidazolate-Cu system as a whole the crystal structure reported last year indicated that the metals were linked by a common histidine residue. There is an exchange interaction between the cobalt and copper however, this is abolished when the linking imidazole is protonated. Further evidence for the close proximity and interactive dependence of the zinc and copper binding sites was obtained from a study of the 4 Cu protein a two-fold enhancement of the activity of 2 Cu dismutase was observed upon occupation of the zinc sites by the Cu ". On the basis of C1 n.m.r. studies, Fee and Ward have suggested that one co-ordination position of Cu in superoxide dismutase is normally occupied by water they further suggest that superoxide can displace the solvent to form a cupric peroxide complex. 

The kinetics of formation of the intermediate complex between catalase and HgOa have been re-examined, and reports of pulse radiolysis experiments with superoxide dismutase and its manganese-containing form have appeared. Bovine superoxide dismutase is known to contain two Cu and two Zn atoms per molecule and it has been shown that the copper site, which is directly involved in the catalytic activity, comprises three nitrogens and a water molecule bound to the metal in a field with less than axial symmetry. Less is known of the zinc site but it has recently been shown, by e.p.r. spectroscopy with the cobalt-copper form of the enzyme, that the... 

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