Electron Paramagnetic Resonance Kinetic Studies of the S States in Spinach Thylakoids

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Abstract
The Tyrz+ decay kinetics have been analyzed by using time-resolved EPR to determine the half-time of each Si → S(i+1) transition in the O2-evolving complex of spinach thylakoids under physiological conditions. Using dark-adapted thylakoids and appropriate single-turnover flash sequences, we were able to detect the signal IIvf kinetics of the Tyrz+ S0 → Tyrz S1, Tyrz+ S1 → Tyrz S2, Tyrz+ S2 → Tyrz S3, and Tyrz+ S3 → (S4) → Tyrz S0 transitions. To correct for damping of the S state synchronization during the flash sequence, the Kok parameters were estimated by measuring the oxygen flash pattern in situ using nitroxide-based EPR oximetry. Following deconvolution of the individual S state contributions, the signal IIvf decay kinetics yield the following half-times for the S state transitions: S0 → S1 in 40−60 μs, S1 → S2 in 85 μs, S2 → S3 in 140 μs, and S3 → (S4) → S0 in 750 μs. Preliminary results with detergent-solubilized PSII membranes suggest that the S3 → S0 transition at least is slowed by a factor of ∼2 in this system. Ramifications of these half-times in terms of electron transfer events on the donor site of PSII are discussed.