MECHANISM OF MN-II OXIDATION BY THE EXTRINSIC 33 KDA PROTEIN OF PHOTOSYSTEM-II

The isolated extrinsic 33 kDa of PSII catalyses the oxidation of Mn-II in the presence of Cl- and a thiol reagent such as beta-mercaptoethan ol. The chloride requirement was specific and the K-m for Cl- was 20 m M. The known inhibitors of O-2 evolution such as NH2OH and NH4+ were s trong inhibitors of this oxidase activity. The reduced form of the 33 kDa protein was active as the oxidase. Addition of P-ME abruptly inhib ited O, evolution in PSII particles initiating an O-2 uptake, suggesti ng that the disulfide form of MSP was essential for O-2 evolution acti vity. While the reduction of the 33 kDa protein by P-ME was shown to b e essential for the induction of Mn-II oxidation to Mn-III associated with O-2 uptake, there was 1:2 stoichiometry between O-2 uptake and Mn -III formed and SH disappearance. The geometry of the catalytic sulfhy dryl site has been probed by fluorometry using O-phthalaldehyde (OPA). A characteristic 337 nm absorption band observed in OPA-bound protein indicated the formation of thioisoindole derivative at the Mn-II bond ing site. The CD spectrum and the fluorescence emission and excitation spectra suggest conformational changes in the oxidized and reduced fo rm of this protein. The results of these studies suggest that tryptoph an 241 and two lysine residues of this protein are in close proximity to functional cysteine residues at the active site. In the proposed re action mechanism for Mn-II oxidase activity, the reduced 33 kDa protei n in the presence of Cl- ions catalyses the oxidation of Mn-II to Mn-I II associated with oxygen uptake.