High-Frequency Electron Nuclear Double-Resonance Spectroscopy Studies of the Mechanism of Proton-Coupled Electron Transfer at the Tyrosine-D Residue of Photosystem II

The solar water-splitting protein complex, photosystem II, catalyzes one of the most energetically demanding reactions in Nature by using light energy to drive the catalytic oxidation of water. Photosystem II contains two symmetrically placed tyrosine residues, Y-D and Y-Z, one on each subunit of the heterodimeric core. The Y-Z residue is kinetically competent and is proposed to be directly involved in the proton-coupled electron transfer reactions of water oxidation. In contrast, the Y-D proton-coupled electron transfer redox poises the catalytic tetranuclear manganese cluster and may electrostatically tune the adjacent monomeric redox-active chlorophyll and beta-carotene in the secondary electron transfer pathway of photosystem II. In this study, we apply pulsed high-frequency electron paramagnetic resonance (EPR) and electron nuclear double-resonance (ENDOR) spectroscopy to study the photochemical proton-coupled electron transfer (PCET) intermediates of Y-D. We detect the "unrelaxed" and "relaxed" photoinduced PCET intermediates of Y-D using high-frequency EPR spectroscopy and observe an increase of the g anisotropy upon temperature-induced relaxation of the unrelaxed intermediate to the relaxed state as previously observed by Faller et al. [(2002) Biochemistry 41, 12914-12920; (2003) Proc. Natl. Acad. Sci. U.S.A. 100, 8732-8735]. This observation suggests the presence of structural differences between the two intermediates. We probe the possible structural differences by performing high-frequency H-2 ENDOR spectroscopy experiments. On the basis of numerical simulations of the experimental H-2 ENDOR spectra, we confirm that (i) there is a significant change in the H-bond length of the tyrosyl radical in the unrelaxed (1.49 angstrom) and relaxed (1.75 angstrom) PCET intermediates. This observation suggests that the D2-His189 residue is deprotonated prior to electron transfer at the Y-D residue and (ii) there are negligible changes in the conformation of the tyrosyl ring in the unrelaxed and relaxed PCET intermediates of Y-D.