Electron transfer properties of the R2 protein of @entity2151 reductase from XXXX .

@entity2769 @entity68 @entity2151 @entity73210 @entity18481 @entity526
The enzyme @entity2151 reductase from @entity526 consists of two proteins, R1 and R2. The active R2 protein contains two dinuclear @entity68 centers and the catalytically essential @entity18481 radical. We have explored the redox properties of the @entity18481 radical and estimate an apparent redox potential of +1000 +/- 100 mV (vs SHE) on the basis of the behavior of numerous mediators. The inability of most of these mediators to equilibrate with the @entity18481 radical supports the notion that the radical exists in an extremely protected hydrophobic pocket that prevents most radical scavengers from interacting with the radical, resulting in its unusual stability. The formal midpoint potential of the diiron clusters of the R2 protein was determined to be -115 +/- 2 mV at pH 7.6 and 4 degrees C. This reduction is a two-electron transfer process, making the R2 protein the first of the nonheme diiron proteins not to stabilize a mixed valence intermediate at ambient temperature. The formal midpoint potential of the dinuclear @entity68 centers is pH dependent, exhibiting a 30 mV/pH unit variance, which indicates that one proton is accepted from the solvent per two electrons transferred to the dinuclear @entity68 center upon reduction. The midpoint potential of the site-directed mutant @entity73210 R2 protein was also investigated under the same conditions, and this midpoint potential was determined to be -178 mV, providing the first direct evidence that the presence of the Y122 residue modulates the redox properties of the diiron clusters. The redox potentials of both the wild type and @entity73210 proteins experience cathodic shifts when measured in the presence of @entity2769 or the R1 protein. For the latter, the midpoint potentials were determined to be -226 mV for the wild type protein and -281 mV for the @entity73210 mutant protein, representing a negative shift of over 100 mV for both proteins. These results indicate that the presence of the Y122 residue does not influence the effect of R1 binding, that the R1 protein preferentially binds the oxidized form of R2, and that the binding of R1 acts as a regulatory control mechanism to prevent unnecessary turnover of the dinuclear @entity68 centers.
