scholarly journals Reduction and inactivation of superoxide dismutase by hydrogen peroxide

1974 ◽  
Vol 139 (1) ◽  
pp. 43-48 ◽  
Author(s):  
Robert C. Bray ◽  
Stephen A. Cockle ◽  
E. Martin Fielden ◽  
Peter B. Roberts ◽  
Giuseppe Rotilio ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Prolonged Exposure ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Limited Range ◽  
Enzymic Activity ◽  
Resonance Spectroscopy ◽  
Paramagnetic Resonance ◽  
Visible Spectra ◽  
Electron Reduction ◽  
Electron Paramagnetic

Reactions of H2O2 with superoxide dismutase were studied by e.p.r. (electron paramagnetic resonance) spectroscopy and other methods. In agreement with earlier work, the Cu2+ of the enzyme is reduced by H2O2, although the reaction does not go to completion and its kinetics are not simple. With dilute enzyme the time for half-reduction with 9mm-H2O2 is about 150ms. It is suggested that the reaction is a one-electron reduction, involving liberation of O2−. On somewhat more prolonged exposure to H2O2, the enzyme is inactivated. For enzyme in dilute solution and over a limited range of H2O2 concentrations, inactivation is first-order with respect to enzyme and reagent, with k=3.1m−1·s−1 at 20–25°C. Inactivation is accompanied by marked changes in the e.p.r. and visible spectra and appears to be associated with destruction of one histidine residue per subunit. It is suggested that this histidine is close to the metal in the native enzyme and essential for its enzymic activity.

scholarly journals Studies by electron-paramagnetic-resonance spectroscopy of the molybdenum centre of spinach (Spinacia oleracea) nitrate reductase

1983 ◽  
Vol 213 (1) ◽  
pp. 137-142 ◽  
Author(s):  
S Gutteridge ◽  
R C Bray ◽  
B A Notton ◽  
R J Fido ◽  
E J Hewitt
Keyword(s):  
Nitrate Reductase ◽  
Prolonged Exposure ◽  
Spinacia Oleracea ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Reaction Times ◽  
Resonance Spectroscopy ◽  
Paramagnetic Resonance ◽  
Sulphite Oxidase ◽  
Electron Paramagnetic ◽  
Reduced Forms

The molybdenum centre of spinach (Spinacia oleracea) nitrate reductase has been investigated by e.p.r. spectroscopy of molybdenum(V) in reduced forms of the enzyme. The resting enzyme gives no signals attributable to Mo(V). However, on reduction with NADH, Mo(V) signals appeared at relatively short reaction times but decreased again on prolonged exposure to excess of the substrate as the enzyme was further reduced. On brief treatment of such samples with nitrate, Mo(V) signals reappeared but disappeared again on longer exposure to excess nitrate as the enzyme became fully reoxidized. Detailed investigation of the signals carried out in both 1H2O and 2H2O revealed the presence of two signal-giving species, referred to as ‘signal A’ and ‘signal B’, analogous to corresponding signals from nitrate reductase from Escherichia coli and from liver sulphite oxidase. Signal A has gav. 1.9767 and shows coupling to a single proton, exchangeable with the solvent, with A(1H)av. 1.3mT, whereas signal B shows no more than weak coupling to protons. Investigation of interconversion between the two species indicated that decreasing the pH from 8.0 to 6.7 had little effect, but that signal A was favoured by the presence of Cl-. This suggests, by analogy with recent work on sulphite oxidase by Bray, Gutteridge, Lamy & Wilkinson [Biochem. J. (1983) 211, 227-236] that Cl- is a ligand of molybdenum in the species giving signal A.

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