A Synechocystis 6803 mutant carrying a chimaeric photosystem II (PSII), in which the Zea mays PsbH subunit (7.7 kDa calculated molecular mass) replaces the cyanobacterial copy (7.0 kDa), was constructed. With the exception of the N-terminal 12 amino acid extension, which has a phosphorylatable threonine, the eukaryotic polypeptide is 78% homologous to its bacterial counterpart. Biochemical characterization of this mutant shows that it expresses the engineered gene correctly and is competent for photoautotrophic growth. Fluorescence analysis and oxygen evolution measurements in the presence of exogenous acceptors indicate that the observed phenotype results from a chimaeric PSII rather than from the absence of function associated with PsbH, suggesting that the heterologous protein is assembled into a functional PSII. Inhibition of oxygen evolution by herbicides belonging to different classes shows that the sensitivity of the mutant PSII is changed only towards phenolic compounds. This result indicates slight conformational modification of the Q B /herbicide binding pocket of the D1 polypeptide caused by the bulky PsbH protein in the mutant, and also suggests close structural interaction of the D1 and PsbH subunits in the topological arrangement of PSII.

The effects of ultraviolet-B radiation (280-320 nm) on photosystem I1 of $JneChoCJJStk sp. PCC 6803 were investigated at the functional and structural levels. Loss of oxygen-evolving and electrontransport activity, measured by various techniques including Clark electrode polarography, fluorescence induction and fluorescence relaxation after a single turnover flash, are discussed in terms of two types of damage caused by ultraviolet-B radiation : (a) depletion of the plastoquinone pool ; (b) perturbation and degradation of the D1 protein, with cleavage in the second transmembrane segment. These findings are in full agreement with those obtained, both in vivo and in vitro for higher plants for which a donor-side mechanism involving the water-splitting Mn cluster has been proposed for the main cleavage of the D1 protein. At the structural level, complete disruption of the photosystem 11 core is documented as a consequence of (or in parallel with) degradation of the D1 protein. From this point of view, ultraviolet-Binduced photoinhibition is unlike the visible-induced type and less susceptible to repair by synthesis and reinsertion of new D1 protein Keywords: photosystem 11; cyanobacteria; ultraviolet-B ; damage; D1 protein.Light energy drives photosynthesis, but excess light is potentially harmful to the photosynthetic apparatus of oxygen-evolving organisms [I]. Numerous studies have investigated the molecular mechanisms underlying photoinactivation of the electron transport activity of photosystem 11 (PSII), which is the main target of photoinhibition, and subsequent degradation of PSI1 reaction centre polypeptides D1 and D2. The quite consensual picture which has emerged up to now is that the reaction centre (RC) acceptor side is initially affected during photoinhibition : non-physiological double reduction of first stable acceptor QA promotes its dissociation from the reaction centre D2 protein, thus irreversibly interrupting the electron transport chain [2]. Further excitation of P680 induces the formation of 'P680 through charge recombination of the radical pair P680+-pheophytin- [3]. The reaction of 'P680 with molecular oxygen produces '0, [ 3 ] , which is thought to cause oxidative damage to the D1 protein, thereby promoting its turnover 14, 51. Following inactivation of the acceptor side, or alternatively, inactivation of RC donor side reactions leads to the increased lifetime of highly oxidizing cation radicals such as P680+, Tyr' and Chl' which are thought to be able to damage the D1 protein [6]. It has been shown that damage to the acceptor and donor sides is associated with different degradation patterns of the D1 protein [7, 81. Abbreviations. CIJnd, 2,6-dichloroindophenol; Chl, chlorophyll ; CP43 and CP47, chlorophyll-protein components of the photosystem TI core: Deriphat, lauryl P-D-in~inopropionidate; QA, first stable electron acceptor in photosystem 11; QR, second stable electron acceptor of photosystem 11; PQ, plastoquinoiie-9; PSII, photosystem 11; RC, reaction centre ; STF, single turnover flas...

No abstract