FTIR detection of water reactions in the oxygen-evolving centre of photosystem II

Flash-induced Fourier transform infrared (FTIR) difference spectroscopy has been used to study the water-oxidizing reactions in the oxygen-evolving centre of photosystem II. Reactions of water molecules were directly monitored by detecting the OH stretching bands of weakly H-bonded OH of water in the 3700–3500 cm −1 region in FTIR difference spectra during S-state cycling. In the S 1 →S 2 transition, a band shift from 3588 to 3617 cm −1 was observed, indicative of a weakened H-bond. Decoupling experiments using D 2 O : H 2 O (1 : 1) showed that this OH arose from a water molecule with an asymmetric H-bonding structure and this asymmetry became more significant upon S 2 formation. In the S 2 →S 3 , S 3 →S 0 and S 0 →S 1 transitions, negative bands were observed at 3634, 3621 and 3612 cm −1 , respectively, representing formation of a strong H-bond or a proton release reaction. In addition, using complex spectral features in the carboxylate stretching region (1600–1300 cm −1 ) as ‘fingerprints’ of individual S-state transitions, pH dependency of the transition efficiencies and the effect of dehydration were examined to obtain the information of proton release and water insertion steps in the S-state cycle. Low-pH inhibition of the S 2 →S 3 , S 3 →S 0 and S 0 →S 1 transitions was consistent with a view that protons are released in the three transitions other than S 1 →S 2 , while relatively high susceptibility to dehydration in the S 2 →S 3 and S 3 →S 0 transitions suggested the insertion of substrate water into the system during these transitions. Thus, a possible mechanism of water oxidation to explain the FTIR data is proposed.