PHOTOPRODUCTION OF THE AZIDYL RADICAL FROM THE AZIDE ANION ON THE OXIDIZING SIDE OF PHOTOSYSTEM-II AND SUPPRESSION OF PHOTOOXIDATION OF TYROSINE-Z BY THE AZIDYL RADICAL

Azide ions inhibited O-2 evolution in PSII membranes from spinach in a time-dependent manner in the light until all activity disappeared. Illumination in the presence of azide (azide-phototreatment) irreversibly inhibited the following processes: (1) both the oxidation of water and the electron transport between the redox-active tyrosine 161 of the D1 protein (Y-Z) and the secondary quinone electron acceptor (Q(B)) site, to the same extent; (2) the donation of electrons to the primary quinone electron acceptor (Q(A)), as measured by monitoring the maximum variable fluorescence of Chl; and (3) the photoproduction of the Y-Z radical (Y-Z(.)). Thus, the primary site of inhibition appeared to lie between Y-Z and Q(A). On illumination of Tris-treated PSII membranes in the presence of azide, production of the azidyl radical was observed by spin-trapping ESR. Yield of Y-Z(.) in Tris-treated membranes on illumination was suppressed by azide. Electron transport from Y-Z to Q(B) in Tris-treated membranes was inhibited only when the azidyl radical was photoproduced, and it was inhibited more rapidly than it was in the oxygenic PSII membranes. These results indicate that the azidyl radical was produced via a univalent oxidation of azide by Y-Z(.) and that it irreversibly inhibited the electron transport from Y-Z to Q(A) in Tris-treated membranes. Although the azidyl radical was undetectable in the oxygenic PSII membranes, probably due to steric interference by the peripheral proteins of water-oxidizing complex with the access of the spin-trapping reagent to the production site of the radical, the participation of the azidyl radical in the inhibition of the oxygenic PSII membranes is suggested since simultaneous occurrence of both electron transport and azide was required for the inhibition. Possible inhibitory mechanisms and the target sites of azidyl radical are discussed.