Direct quantification of the four individual S states in Photosystem II using EPR spectroscopy

EPR spectroscopy is very useful in studies of the oxygen evolving cycle in Photosystem II and EPR signals from the CaMn 4 cluster are known in all S states except S 4. Many signals are insufficiently understood and the S 0, S 1, and S 3 states have not yet been quantifiable through their EPR signals. Recently, split EPR signals, induced by illumination at liquid helium temperatures, have been reported in the S 0, S 1, and S 3 states. These split signals provide new spectral probes to the S state chemistry. We have studied the flash power dependence of the S state turnover in Photosystem II membranes by monitoring the split S 0, split S 1, split S 3 and S 2 state multiline EPR signals. We demonstrate that quantification of the S 1, S 3 and S 0 states, using the split EPR signals, is indeed possible in samples with mixed S state composition. The amplitudes of all three split EPR signals are linearly correlated to the concentration of the respective S state. We also show that the S 1 → S 2 transition proceeds without misses following a saturating flash at 1 °C, whilst substantial misses occur in the S 2 → S 3 transition following the second flash.