Location of the calcium binding site in Photosystem II: A Mn2+ substitution study.

Abstract

The whereabouts of the Ca²⁺ site in Photosystem II (PSII) was investigated by experiments in which Mn²⁺ was substituted for Ca²⁺. When stoichiometric amounts of Mn²⁺ ions were added to Ca²⁺-depleted PSII, the Mn²⁺ was not detected by EPR. The titration of Ca²⁺ back into Ca²⁺-depleted/Mn²⁺-containing PSII resulted in the simultaneous release of the Mn²⁺ and the loss of the two EPR signals which are characteristic of the Ca²⁺-depleted enzyme (i.e., the stable, modified S₂ multiline signal arising from the intrinsic Mn cluster and the split S₃ signal from an organic radical interacting with the Mn cluster). These results indicate that the Mn²⁺ occupies the functional Ca²⁺ site. The S₂ and S₃ EPR signal characteristic of this kind of Ca²⁺-depleted preparation were unaffected by the binding of the Mn²⁺ Since, from earlier results, it seems likely that the modification and stability of S₂ multiline signal in these PSII preparations is due to binding of chelator to or close to the Mn cluster, the present results indicate that the Ca²⁺ site (at least when occupied by Mn²⁺) does not overlap with the chelator binding site. Since Mn²⁺ binding does not effect the S₂ EPR signal from the Mn cluster, it can be concluded that the Mn²⁺ is not involved in detectable magnetic interactions with the cluster. This result indicates that the Mn²⁺-occupied Ca²⁺ binding site is outside the first co-ordination sphere of the Mn cluster. The relaxation properties of TyrD^. were enhanced by the presence of the Mn²⁺ when the Mn cluster was in the S₁ state.