Mutations in the CD-loop region of the D2 protein in Synechocystis sp PCC 6803 modify charge recombination pathways in photosystem II in vivo

The lumenal CD-loop region of the D2 protein of photosystem II contains res idues that interact with the primary electron donor P680 and the redox acti ve tyrosyl residue Y-D. Photosystem II properties were studied in a number of photoautotrophic mutants of Synechocystis sp. PCC 6803, most of which ca rried combinatorial mutations in residues 164-170, 179-186, or 187-194 of t he D2 protein. To facilitate characterization of photosystem II properties in the mutants, the CD-loop mutations were introduced into a photosystem I- less background. According to variable fluorescence decay measurements in D CMU-treated cells, charge recombination of Q(A)(-) with the donor side was faster in the majority of mutants (t(1/2) = 45-140 ms) than in the control (t(1/2) = 180 ms). However, in one mutant (named C7-3), the decay of Q(A)(- ) was 2 times slower than in the control (t(1/2) = 360 ms). The decay half- time of each mutant correlated with the yield of the Q-band of thermolumine scence (TL) emitted due to S(2)Q(A)(-) charge recombination. The C7-3 mutan t had the highest TL intensity, whereas no Q-band was detected in the mutan ts with fast Q(A)(-) decay (t(1/2) = 45-50 ms). The correlated changes in t he rate of recombination and in TL yield in these strains suggest the exist ence of a nonradiative pathway of charge recombination between Q(A)(-) and the donor side. This may involve direct electron transfer from Q(A)(-) to P 680(+) in a way not leading to formation of excited chlorophyll. Many mutat ions in the CD-loop appear to increase the equilibrium P680(+) concentratio n during the lifetime of the S(2)Q(A)(-) state, for example, by making the midpoint potential of the P680(+)/P680 redox couple more negative. The nonr adiative charge recombination pathway involves a low activation energy and is less temperature-dependent than the formation of excited P680 that leads to TL emission. Therefore, during the TL measurements in these mutants, th e S(2)Q(A)(-) State can recombine nonradiatively before temperatures are It ached at which radiative charge recombination becomes feasible. The result s presented here highlight the presence of two charge recombination pathway s and the importance of the CD-loop of the D2 protein in determination of t he energy gap between the P680(+)S(1) and P680S(2) states.