Site-directed mutagenesis of basic arginine residues 305 and 342 in the CP43 protein of photosystem II affects oxygen-evolving activity in Synechocystis 6803

The intrinsic chlorophyll protein CP 43, a component of photosystem II (PS II) in higher plants, green algae, and cyanobacteria, is encoded by the psb C gene. Oligonucleotide-directed mutagenesis was employed to introduce muta tions into a segment of psbC that encodes the large extrinsic loop E of CP 43 in the cyanobacterium Synechocystis 6803. Two mutations, R305S and R342S , each produced a strain with impaired photosystem II activity. The R305S m utant strain grew photoautotrophically at rates comparable to the control s train, Immunological analyses of a number of PSII components indicated that this mutant accumulated. normal quantities of PSII proteins. However, this mutant evolved oxygen to only 70% of control rates at saturating Tight int ensifies. Measurements of total variable fluorescence yield indicated that this mutant assembled approximately 70% of the PSII centers found in the co ntrol strain. The R342S mutant failed to grow photoautotrophically and exhi bited no capacity for oxygen evolution. However, when grown photoheterotrop hically in medium containing both glucose and 3-(3,4-dichlorophenyl)-1,1-di methylurea (DCMU), oxygen-evolving activity was observed in the R342S mutan t, but at a low level of approximately 10% of the control rate. Immunologic al analysis of isolated thylakoid membranes from this mutant also indicated that this strain accumulated normal amounts of PSII core proteins. Total v ariable fluorescence yields for the R342S mutant indicated that it assemble d a severely reduced number of fully functional PSII centers. R305S and R34 2S mutant strains exhibited, respectively, 2.7- and 4-fold increased sensit ivity to photoinactivation. The fluorescence rise times for both mutants we re comparable to the control when hydroxylamine was used as electron donor. However, both strains exhibited an increase (2.5- and 8-fold, respectively , for R305S and R342S) in fluorescence rise times with water as an electron donor. These results suggest that the mutations R305S and R342S each produ ce a defect associated with the oxygen-evolving complex of photosystem II. These are the first site-directed mutations in CP 43 to show such an effect .