Electrostatic control of the rate-determining step of the copper, zinc superoxide dismutase catalytic reaction.
The dependence of the activity of bovine Cu,Zn superoxide dismutase on pH and ionic strength was extensively investigated in the ranges of pH 7.4-pH 12.3 and of ionic strength of 0.02-0.25 M. The results obtained indicate that two positively charged groups having pK values of approximately 10.1 and 10.8 are involved in the control of the activity. On the basis of previous work on the three-dimensional structure and on the chemically modified enzyme, these groups are likely to be lysine side chains, in particular Lys-120 and Lys-134. The oxidation state of the enzyme-bound copper ion at the steady state was found to be the same at either pH 7.4 or pH 11.5. The diffusion of superoxide ion into the active site, which is controlled by the positive charges around the active site itself, appears to be the rate-determining step of the dismutation reaction. NMR measurements of the relaxation rates of F- showed that this control also applies to the access of F- to the active site. Comparison of the nuclear relaxation rates of F- with the enzyme activity indicates that F- relaxation is controlled by the deprotonation of the group with pK approximately 10.8, which appears to be responsible for about 50% of the total activity measured at neutral pH.
D000818:Animals; D002417:Cattle; D004563:Electrochemistry; D004912:Erythrocytes / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D009994:Osmolar Concentration; D013482:Superoxide Dismutase / Q000097:blood*
D013482:Superoxide Dismutase