Molecular origin of the pH dependence of tyrosine D oxidation kinetics and radical stability in photosystem II

Abstract

A role for redox-active tyrosines has been demonstrated in many important biological processes, including water oxidation carried out by photosystem II (PSII) of oxygenic photosynthesis. The rates of tyrosine oxidation and reduction and the Tyr /Tyr reduction potential are undoubtedly controlled by the immediate environment of the tyrosine, with the coupling of electron and proton transfer, a critical component of the kinetic and redox behavior. It has been demonstrated by Faller et al. that the rate of oxidation of tyrosine D (Tyr D) at room temperature and the extent of Tyr D oxidation at cryogenic temperatures, following flash excitation, dramatically increase as a function of pH with a p K a of ≈ 7.6 [Faller et al. 2001 Proc. Natl. Acad. Sci. USA 98, 14368–14373; Faller et al. 2001 Biochemistry 41, 12914–12920]. In this work, we investigated, using FTIR difference spectroscopy, the mechanistic reasons behind this large pH dependence. These studies were carried out on Mn-depleted PSII core complexes isolated from Synechocystis sp. PCC 6803, WT unlabeled and labeled with 13C 6-, or 13C 1(4)-labeled tyrosine, as well as on the D2-Gln164Glu mutant. The main conclusions of this work are that the pH-induced changes involve the reduced Tyr D state and not the oxidized Tyr D state and that Tyr D does not exist in the tyrosinate form between pH 6 and 10. We can also exclude a change in the protonation state of D2-His189 as being responsible for the large pH dependence of Tyr D oxidation. Indeed, our data are consistent with D2-His189 being neutral both in the Tyr D and Tyr D states in the whole pH6-10 range. We show that the interactions between reduced Tyr D and D2-His189 are modulated by the pH. At pH greater than 7.5, the ν(CO) mode frequency of Tyr D indicates that Tyr D is involved in a strong hydrogen bond, as a hydrogen bond donor only, in a fraction of the PSII centers. At pH below 7.5, the hydrogen-bonding interaction formed by Tyr D is weaker and Tyr D could be also involved as a hydrogen bond acceptor, according to calculations performed by Takahashi and Noguchi [J. Phys. Chem. B 2007 111, 13833–13844]. The involvement of Tyr D in this strong hydrogen-bonding interaction correlates with the ability to oxidize Tyr D at cryogenic temperatures and rapidly at room temperature. A strong hydrogen-bonding interaction is also observed at pH 6 in the D2-Gln164Glu mutant, showing that the residue at position D2-164 regulates the properties of Tyr D. The IR data point to the role of a protonatable group(s) (with a p K a of ≈ 7) other than D2-His189 and Tyr D, in modifying the characteristics of the Tyr D hydrogen-bonding interactions, and hence its oxidation properties. It remains to be determined whether the strong hydrogen-bonding interaction involves D2-His189 and if Tyr D oxidation involves the same proton transfer route at low and at high pH.