Rapid loss of structural motifs in the manganese complex of oxygenic photosynthesis by x-ray irradiation at 10-300 K

Structural changes upon photoreduction caused by x-ray irradiation of the water-oxidizing tetramanganese complex of photosystem II were investigated by x-ray absorption spectroscopy at the manganese K-edge. Photoreduction was directly proportional to the x-ray dose. It was faster in the higher oxidized S-2 state than in S-1; seemingly the oxidizing potential of the metal site governs the rate. X-ray irradiation of the S-1 state at 15 K initially caused single-electron reduction to S-0* accompanied by the conversion of one di-mu-oxo bridge between manganese atoms, previously separated by similar to 2.7 angstrom, to a mono-mu-oxo motif. Thereafter, manganese photoreduction was 100 times slower, and the biphasic increase in its rate between 10 and 300 K with a breakpoint at similar to 200 K suggests that protein dynamics is rate-limiting the radical chemistry. For photoreduction at similar x-ray doses as applied in protein crystallography, halfway to the final Mn-4(II) state the complete loss of inter-manganese distances < 3 angstrom was observed, even at 10 K, because of the destruction of mu-oxo bridges between manganese ions. These results put into question some structural attributions from recent protein crystallography data on photosystem II. It is proposed to employ controlled x-ray photoreduction in metalloprotein research for: (i) population of distinct reduced states, (ii) estimating the redox potential of buried metal centers, and (iii) research on protein dynamics.