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 CaMn4 cluster are known in all S states except S4. Many signals are insufficiently understood and the S0, S1, and S3 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 S0, S1, and S3 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 S0, split S1, split S3 and S2 state multiline EPR signals. We demonstrate that quantification of the S1, S3 and S0 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 S1 → S2 transition proceeds without misses following a saturating flash at 1 °C, whilst substantial misses occur in the S2 → S3 transition following the second flash.