Electron paramagnetic resonance kinetic studies of the S states in spinach thylakoids

The Tyr(2)(+) decay kinetics have been analyzed by using time-resolved EPR to determine the half-time of each S-i --> S-(i+l) transition in the O-2-evolving complex of spinach thylakoids under physiological conditions. Using dark-adapted thylakoids and appropriate single-turnover flash sequences, we wen able to detect the signal IIvf kinetics of the Tyr(z)(+) S-0 --> Tyr(z)(+) S-1, Tyr(z)(+) S-1 --> Tyr(z) S-2, Tyr(z)(+) S-2 --> Tyr(z) S-3, and Tyr(z)(+) S-3 --> (S-4) --> Tyr(z) S-0 transitions. To correct for damping of the S state synchronization during the Rash 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 slate contributions, the signal IIvf decay kinetics yield the following half-times for the S state transitions: S-0 --> S-1 in 40-60 mu s, S-1 --> S-2 in 85 mu s, S-2 --> S-3 in 140 mu s, and S-3 --> (S-4) --> S-0 in 750 mu s. Preliminary results with detergent-solubilized PSII membranes suggest that the S-3 --> S-0 transition at least is slowed by a factor of similar to 2 in this system. Ramifications of these half-times in terms of electron transfer events on the donor site of PSII are discussed.