title: Control of the evolution of iron peroxide intermediate in superoxide reductase from Desulfoarculus baarsii. Involvement of lysine 48 in protonation.
abstract: Superoxide reductase is a nonheme iron metalloenzyme that detoxifies superoxide anion radicals O(2)(•-) in some microorganisms. Its catalytic mechanism was previously proposed to involve a single ferric iron (hydro)peroxo intermediate, which is protonated to form the reaction product H(2)O(2). Here, we show by pulse radiolysis that the mutation of the well-conserved lysine 48 into isoleucine in the SOR from  Desulfoarculus baarsii  dramatically affects its reaction with O(2)(•-). Although the first reaction intermediate and its decay are not affected by the mutation, H(2)O(2) is no longer the reaction product. In addition, in contrast to the wild-type SOR, the lysine mutant catalyzes a two-electron oxidation of an olefin into epoxide in the presence of H(2)O(2), suggesting the formation of iron-oxo intermediate species in this mutant. In agreement with the recent X-ray structures of the peroxide intermediates trapped in a SOR crystal, these data support the involvement of lysine 48 in the specific protonation of the proximal oxygen of the peroxide intermediate to generate H(2)O(2), thus avoiding formation of iron-oxo species, as is observed in cytochrome P450. In addition, we proposed that the first reaction intermediate observed by pulse radiolysis is a ferrous-iron superoxo species, in agreement with TD-DFT calculations of the absorption spectrum of this intermediate. A new reaction scheme for the catalytical mechanism of SOR with O(2)(•-) is presented in which ferrous iron-superoxo and ferric hydroperoxide species are reaction intermediates, and the lysine 48 plays a key role in the control of the evolution of iron peroxide intermediate to form H(2)O(2).
authors.affiliation: CEA, iRTSV, Laboratoire de Chimie et Biologie des Métaux, F-38054 Grenoble, France.
authors.forename: Florence
authors.initials: F
authors.lastname: Bonnot
authors.forename: Thibaut
authors.initials: T
authors.lastname: Molle
authors.forename: Stéphane
authors.initials: S
authors.lastname: Ménage
authors.forename: Yohann
authors.initials: Y
authors.lastname: Moreau
authors.forename: Simon
authors.initials: S
authors.lastname: Duval
authors.forename: Vincent
authors.initials: V
authors.lastname: Favaudon
authors.forename: Chantal
authors.initials: C
authors.lastname: Houée-Levin
authors.forename: Vincent
authors.initials: V
authors.lastname: Nivière
mesh_terms: D020564:Deltaproteobacteria / Q000201:enzymology*; D019143:Evolution, Molecular; D005290:Ferric Compounds / Q000737:chemistry / Q000378:metabolism*; D008239:Lysine / Q000737:chemistry / Q000378:metabolism*; D010088:Oxidoreductases / Q000737:chemistry / Q000378:metabolism*; D010545:Peroxides / Q000737:chemistry / Q000378:metabolism*; D011522:Protons; D011789:Quantum Theory
chemical_list: D005290:Ferric Compounds; D010545:Peroxides; D011522:Protons; D010088:Oxidoreductases; C121849:superoxide reductase; D008239:Lysine