Effects of pH on the S3 State of the Oxygen Evolving Complex in Photosystem II Probed by EPR Split Signal Induction

The electrons extracted from the CaMn4 cluster during water oxidation in photosystem II are transferred to P680 + via the redox-active tyrosine D1-Tyr161 (YZ). Upon YZ oxidation a proton moves in a hydrogen bond toward D1-His190 (HisZ). The deprotonation and reprotonation mechanism of YZ-OH/YZ-O is of key importance for the catalytic turnover of photosystem II. By light illumination at liquid helium temperatures (∼5 K) YZ can be oxidized to its neutral radical, YZ •. This can be followed by the induction of a split EPR signal from YZ • in a magnetic interaction with the CaMn4 cluster, offering a way to probe for YZ oxidation in active photosystem II. In the S3 state, light in the near-infrared region induces the split S3 EPR signal, S2′YZ •. Here we report on the pH dependence for the induction of S2′YZ • between pH 4.0 and pH 8.7. At acidic pH the split S3 EPR signal decreases with the apparent pK a (pK app) ∼ 4.1. This can be correlated to a titration event that disrupts the essential H-bond in the YZ-HisZ motif. At alkaline pH, the split S3 EPR signal decreases with the pK app ∼ 7.5. The analysis of this pH dependence is complicated by the presence of an alkaline-induced split EPR signal (pK app ∼ 8.3) promoted by a change in the redox potential of YZ. Our results allow dissection of the proton-coupled electron transfer reactions in the S3 state and provide further evidence that the radical involved in the split EPR signals is indeed YZ •.