Probing reactive sites within the Photosystem II manganese cluster: Evidence for separate populations of manganese that differ in redox potential

When the oxygen evolving complex of Photosystem II is depleted of essential cofactor atoms (Ca2+ and Cl-) by a high ionic strength treatment that extracts 23 and 17 kDa extrinsic polypeptides, Mn2+ can be released by several large reductants (hydroquinone (H(2)Q), N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD), methyl derivatives of hydroxylamine). This reactivity can be slowed if the enzyme is reconstituted with Ca2+. For TMPD, data from EPR and X-ray absorption spectroscopy indicate that Ca2+ reconstitution restricts initial Mn reduction to a site which contains a Mn4+ atom. Dimethylhydroxylamine (DMHA), a much smaller Mn reductant, is unable to reduce the cluster under these same conditions, even though DMHA and TMPD can generate Mn2+ from the Mn cluster in the absence of Ca2+. These reductants differ in redox potential by about 300 mV, and it is likely that the higher potential reductant, DMHA (+550 mV), is restricted to initially reacting with a Mn3+ species that is screened by the Ca2+ atom and is incapable, in the presence of Ca2+ of reducing lower potential atoms of the cluster that do react with TMPD. Reduction of the reactive Mn3+ species by DMHA in the absence of Ca2+, however, subsequently allows reduction of the remaining 3 Mn. Such an arrangement of metals and potentials in the cluster is in accord with models for the site of water oxidation and for the structure of the Ca-Mn-4 cluster.