Jun Minagawa1, D. M. Kramer2, Atsuko Kanazawa1 and A. R. Crofts1,3
1Program in Biophysics, 2Dept. of Plant Biology, and 3Dept. of Microbiol
University of Illinois, 607 S, Mathews Ave., Urbana, IL 61801, USA
The electron donor to oxidized P680, tyrosine Z (Y161) in the D1 polypeptide of photosystem II in C. reinhardtii, was replaced by phenylalanine via site-directed mutagenesis, to produce a pho- strain. We have examined the functional consequences by measuring flash-induced changes in fluorescence yield (f) and absorbance. In the presence of DCMU, f was low after a single flash, indicating that no rapid donor was present; a rise in f on later flashes showed a slow electron donor to accumulated P680+. To identify this donor species, we looked at optical absorbance changes on moderate light illumination. Both chlorophyll(s) and carotenoid(s) were bleached at a rate which depended strongly on experimental conditions. In the absence of O2, or upon addition of DCMU or hydroquinone, carotenoid bleaching was inhibited. This suggests that mutation of the donor Y161 allowed photoaccumulation of oxidized P680, which could be re-reduced by exogenous electron donors, such as hydroquinone, or by endogenous pigments. We are now studying the mechanism of the oxygen dependency of carotenoid bleaching. Additional D1 mutants affecting donor side reactions are N191 to L, D or A, and E189 to L, D, or Q. We have characterized the equilibria on the donor-side in these mutants using studies of the back-reaction, and thermoluminescence.