D1-Asn-298 in photosystem II is involved in a hydrogen-bond network near the redox-active tyrosine Y_Z for proton exit during water oxidation

In photosynthetic water oxidation, two water molecules are converted into one oxygen molecule and four protons at the Mn₄CaO₅ cluster in photosystem II (PSII) via the S-state cycle. Efficient proton exit from the catalytic site to the lumen is essential for this process. However, the exit pathways of individual protons through the PSII proteins remain to be identified. In this study, we examined the involvement of a hydrogen-bond network near the redox-active tyrosine Y_Z in proton transfer during the S-state cycle. We focused on spectroscopic analyses of a site-directed variant of D1-Asn-298, a residue involved in a hydrogen-bond network near Y_Z. We found that the D1-N298A mutant of Synechocystis sp. PCC 6803 exhibits an O₂ evolution activity of 10% of the wild-type. D1-N298A and the wild-type D1 had very similar features of thermoluminescence glow curves and of an FTIR difference spectrum upon Y_Z oxidation, suggesting that the hydrogen-bonded structure of Y_Z and electron transfer from the Mn₄CaO₅ cluster to Y_Z were little affected by substitution. In the D1-N298A mutant, however, the flash-number dependence of delayed luminescence showed a monotonic increase without oscillation, and FTIR difference spectra of the S-state cycle indicated partial and significant inhibition of the S₂→ S₃ and S₃→ S₀ transitions, respectively. These results suggest that the D1-N298A substitution inhibits the proton transfer processes in the S₂ → S₃ and S₃ → S₀ transitions. This in turn indicates that the hydrogen-bond network near Y_Z can be functional as a proton transfer pathway during photosynthetic water oxidation.