No evidence from FTIR difference spectroscopy that aspartate-342 of the D1 polypeptide ligates a Mn ion that undergoes oxidation during the S₀ to S₁, S₁ to S₂, or S₂ to S ₃ transitions in photosystem II

In the recent X-ray crystallographic structural models of photosystem II, Asp342 of the D1 polypeptide is assigned as a ligand of the oxygen-evolving Mn₄ cluster. To determine if D1-Asp342 ligates a Mn ion that undergoes oxidation during one or more of the S₀ → S ₁, S₁ → S₂, and S₂ → S₃ transitions, the FTIR difference spectra of the individual S state transitions in D1-D342N mutant PSII particles from the cyanobacterium Synechocystis sp. PCC 6803 were compared with those in wild-type PSII particles. Remarkably, the data show that the mid-frequency (1800-1200 cm^-1) FTIR difference spectra of wild-type and D1-D342N PSII particles are essentially identical. Importantly, the mutation alters none of the carboxylate vibrational modes that are present in the wild-type spectra. The absence of significant mutation-induced spectral alterations in D1-D342N PSII particles shows that the oxidation of the Mn₄ cluster does not alter the frequencies of the carboxylate stretching modes of D1-Asp342 during the S₀ → S ₁, S₁ → S₂, or S₂ → S ₃ transitions. One explanation of these data is that D1-Asp342 ligates a Mn ion that does not increase its charge or oxidation state during any of these S state transitions. However, because the same conclusion was reached previously for D1-Asp170, and because the recent X-ray crystallographic structural models assign D1-Asp170 and D1-Asp342 as ligating different Mn ions, this explanation requires that (1) the extra positive charge that develops on the Mn₄ cluster during the S₁ → S₂ transition be localized on the Mn ion that is ligated by the α-COO ^- group of D1-Ala344 and (2) any increase in positive charge that develops on the Mn₄ cluster during the S₀ → S ₁ and S₂ → S₃ transitions be localized on the one Mn ion that is not ligated by D1-Asp170, D1-Asp342, or D1-Ala344. In separate experiments that were conducted with L-[1-¹³C]alanine, we found no evidence that D1-Asp342 ligates the same Mn ion that is ligated by the α-COO^- group of D1-Ala344.