Mechanistic insight into O=O bond formation upon model‐independent visualization of the coordination geometry and ligand composition of Mn 4 Ca cofactor in dark‐adapted photosystem II structures

The Mn 4 Ca cofactor of photosystem II (PSII), which is found in its oxygen‐evolving center (OEC), catalyzes the oxidation of water. Spectroscopic studies performed on dark‐adapted PSII samples have led to two mutually incompatible hypotheses about the oxidation states of these manganese ions: Mn(III) 4 or Mn(III) 2 Mn(IV) 2 . It should be possible to determine which is correct crystallographically because they differ in their implications for manganese–ligand bond lengths and coordination geometries. Reported here are the results of a detailed analysis of the electron density in the Mn 4 Ca region of OEC‐omit maps derived from a set of published X‐ray crystal structures of dark‐adapted PSII, the data for which were collected using three different X‐ray doses so that the effects of radiation damage could be assessed. This analysis supports the conclusion that Mn(III) 4 is correct and that all of the Mn(III) ions in the OEC are square‐pyramidally coordinated, i.e. pentadentate. It is further evident that the oxygen ligand sites in these complexes are not fully occupied, and that occupancies vary from one PSII sample to the next. The average occupancies per oxygen ligand site ranged from 0.74 to 0.86 in the set of PSII samples analyzed here. Because fewer than 1% of the OECs were reported as having been damaged in the lowest dose structure, the oxidation number of the manganese ions in X‐ray‐naïve, dark‐adapted OECs would appear to be Mn(III) 4 . The geometry of the protein component of PSII, which controls inter‐manganese distances in the OEC, suggests a reverse four‐step/four‐chamber combustion engine mechanism for O=O bond formation.