Tyrosine radicals are involved in the photosynthetic oxygen-evolving system

In addition to the reaction-center chlorophyll, at least two other organic cofactors are involved in the photosynthetic oxygen-evolution process. One of these cofactors, called "Z," transfers electrons from the site of water oxidation to the reaction center of photosystem II. The other species, "D," has an uncertain function but gives rise to the stable EPR signal known as signal II. Z+. and D+. have identical EPR spectra and are generally assumed to arise from species with the same chemical structure. Results from a variety of experiments have suggested that Z and D are plastoquinones or plastoquinone derivatives. In general, however, the evidence to support this assignment is indirect. To address this situation, we have developed more direct methods to assign the structure of the Z+./D+. radicals. By selective in vivo deuteration of the methyl groups of plastoquinone in cyanobacteria, we show that hyperfine couplings from the methyl protons cannot be responsible for the partially resolved structure seen in the D+. EPR spectrum. That is, we verify by extraction and mass spectrometry that quinones are labeled in algae fed deuterated methionine, but no change is observed in the line shape of signal II. Considering the spectral properties of the D+. radical, a tyrosine origin is a reasonable alternative. In a second series of experiments, we have found that deuteration of tyrosine does indeed narrow the D+. signal. Extraction and mass spectral analysis of the quinones in these cultures show that they are not labeled by tyrosine. These results eliminate a plastoquinone origin for D+.; we conclude instead that D+., and most likely Z+., are tyrosine radicals.