In the preceding paper the mechanism of catalysis of the manganese-containing superoxide dismutase from Bacillus stearothermophilus was shown to involve a ‘fast cycle’ and a ‘slow cycle’ [McAdam, Fox, Lavelle & Fielden, 1977 (Biochem. J. 165, 71-79)]. Further properties of the enzyme was considered in the present paper. Pulse-radiolysis studies, under conditions of low substrate concentration to (i.e. when the fast cycle predominates), showed that enzyme activity decreases as pH increases (6.5-10.2). Activity was unaffected by the addition of H2O2 or NaN3 but slightly decreased by KCN. Both H2O2 and the reducing radical anion CO2– caused a decrease in A480 of the native enzyme. The rate of the fast catalytic cycle was independent of temperature (5-55 degrees C), and as temperature increases the slow cycle becomes relatively more important. Arrhenius parameters of the rate contants were estimated. The possible identity of the various forms of the enzyme is considered.

The enzymic reaction mechanism of a manganese-containing superoxide dismutase from Bacillus stearothermophilus was studied by using pulse radiolysis. During catalysis (pH 8.9; 25 degrees C), changes occurring in the kinetics of substrate disappearance and in the visible absorption of the enzyme at 480 nm established that the simple two-step mechanism found for copper- and iron-containing superoxide dismutases is not involved. At a low ratio (less than 15) of substrate concentration to enzyme concentration the decay of O2–is close to exponetial, whereas at much higher ratios (greater than 100) the observed decay is predominantly zero-order. The simplest interpretation of the results invokes a rapid one-electron oxidation-reduction cycle (‘the fast cycle’) and, concurrently, a slower reaction giving a form of the enzyme that is essentially unreactive towards O2– but which undergoes a first-order decay to yield fully active native enzyme (‘the slow cycle’). The fast cycle involves the native enzyme EA and a form of the enzyme EB which can be obtained also by treating the form EA with H2O2. Computer calculations made with such a simple model predict behaviour in excellent agreement with the observed results.