A pulse-radiolysis study of the manganese-containing superoxide dismutase from Bacillus stearothermophilus. A kinetic model for the enzyme action

Abstract

The enzymic reaction mechanism of a Mn-containing superoxide dismutase from B. stearothermophilus was studied by using pulse radiolysis. During catalysis (pH 8.9; 25.degree. C), changes occurring in the kinetics of substrate disappearance and in the visible absorption of the enzyme at 480 nm established that the simple 2-step mechanism found for Cu- and Fe-containing superoxide dismutases is not involved. At a low ratio (< 15) of substrate concentration to enzyme concentration the decay of .**GRAPHIC**. is close to exponential, but at much higher ratios (> 100) the observed decay is predominantly zero-order. The simplest interpretation of the results invokes a rapid 1-electron oxidation-reduction cycle (the fast cycle) and, concurrently, a slower reaction giving a form of the enzyme that is essentially unreactive towards .**GRAPHIC**. but which undergoes a 1st-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 behavior in excellent agreement with the observed results.