  Superoxide reductase (SOR) is a small metalloenzyme that catalyzes reduction
of O(2)(*)(-) to H(2)O(2) and thus provides an antioxidant mechanism against
superoxide radicals. Its active site contains an unusual mononuclear ferrous
center, which is very efficient during electron transfer to O(2)(*)(-)
[Lombard, M., Fontecave, M., Touati, D., and Nivi{\`e}re, V. (2000) J. Biol.
Chem. 275, 115-121]. The reaction of the enzyme from Desulfoarculus baarsii
with superoxide was studied by pulse radiolysis methods. The first step is an
extremely fast bimolecular reaction of superoxide reductase with superoxide,
with a rate constant of (1.1 +/- 0.3) x 10(9) M(-1) s(-1). A first intermediate
is formed which is converted to a second one at a much slower rate constant of
500 +/- 50 s(-1). Decay of the second intermediate occurs with a rate constant
of 25 +/- 5 s(-1). These intermediates are suggested to be iron-superoxide and
iron-peroxide species. Furthermore, the role of glutamate 47 and lysine 48,
which are the closest charged residues to the vacant sixth iron coordination
site, has been investigated by site-directed mutagenesis. Mutation of glutamate
47 into alanine has no effect on the rates of the reaction. On the contrary,
mutation of lysine 48 into an isoleucine led to a 20-30-fold decrease of the
rate constant of the bimolecular reaction, suggesting that lysine 48 plays an
important role during guiding and binding of superoxide to the iron center II.
In addition, we report that expression of the lysine 48 sor mutant gene hardly
restored to a superoxide dismutase-deficient Escherichia coli mutant the
ability to grow under aerobic conditions.
