| Abstract
| - Superoxide reductase (SOR) catalyzes the reduction of O2•- to H2O2. Its active site consists ofa non-heme Fe2+ center in an unusual square-pyramidal [His4 Cys] coordination. Like many SORs, theelectronic absorption band corresponding to the oxidized active site of the SOR from Desulfoarculusbaarsiiexhibits a pH-dependent alkaline transition changing from ca. 644 to 560 nm as the pH increases and withan apparent pKa of 9.0. Variants in which the conserved amino acids glutamate 47 and lysine 48 werereplaced by the neutral residues alanine (E47A) and isoleucine (K48I), respectively, exhibited the samealkaline transition but at lower apparent pKa values of 6.7 and 7.6, respectively. Previous work [Nivière, V.;Asso, M.; Weill, C. O.; Lombard, M.; Guigliarelli, B.; Favaudon, V.; Houée-Levin, C. Biochemistry2004,43, 808−818] has shown that this alkaline transition is associated with the protonation/deprotonation of anunidentified base, B-, which is neither E47 nor K48. In this work, we show by resonance Ramanspectroscopy that at basic pH a high-spin Fe3+-OH species is formed at the active site. The presence ofthe HO- ligand was directly associated with an absorption band maximum at 560 nm, whereas uponprotonation, the band shifts to 644 nm. With respect to our previous work, B- can be identified with thishigh-spin Fe3+-OH species, which upon protonation results in a water molecule at the active site. Implicationsfor the SOR catalytic cycle are proposed.
|
