Mechanism of Methanol Inhibition of Photosynthetic Water Oxidation As Studied by Fourier Transform Infrared Difference and Time-Resolved Infrared Spectroscopies.

Photosynthetic water oxidation is performed at the Mn4CaO5 cluster in photosystem II. In this study, we investigated the effect of methanol, an analogue of water, on the water oxidation reaction and its interaction site using Fourier transform infrared (FTIR) difference and time-resolved infrared (TRIR) spectroscopies. Flash-induced FTIR difference measurement of the S-state cycle showed that methanol decreases mainly the efficiency of the S3 → S0 transition. TRIR measurement further showed that methanol slowed the rates of the S2 → S3 and S3 → S0 transitions. FTIR difference spectra upon the S1 → S2 transition exhibited prominent methanol-induced changes in the amide I and II bands of the main chains, whereas little change was observed in the bands of carboxylate groups, histidine side chains, and a water network in the vicinity of the Mn4CaO5 cluster. Similar tendencies were also observed with ethanol and 2-propanol. The C-O stretching vibration of methanol was further identified in the S2-minus-S1 spectrum using 18O-labeled methanol. These results indicate that methanol and small alcohols are bound near the Mn4CaO5 cluster but with no direct interaction. They probably replace a water molecule in a water channel around the Mn4CaO5 cluster, possibly interacting with a main chain amide. It is thus suggested that this replacement of water with methanol or a small alcohol inhibits water/proton transfer during the S2 → S3 and S3 → S0 transitions, which in turn provides experimental support for the view that these two transitions involve the water uptake and proton release processes.