Calcium depletion modifies the structure of the photosystem II O(2)-evolving complex.
A 5 min exposure of photosystem II to a pH 3 citric acid solution is a simple method for selective removal of Ca(2+) from the O(2)-evolving complex. The resulting preparation retains the 23 and 17 kDa extrinsic polypeptides, but the activity of this material is only 10-20% of that of an untreated control sample. Biochemical characterization of citrate-treated photosystem II reveals that some reaction centers lose the extrinsic proteins during citrate treatment. Furthermore, a comparison of photosystem II preparations treated with citrate, or depleted of 23 and 17 kDa extrinsic polypeptides by high-salt treatment, shows that low concentrations of a small reductant, NH(2)OH, which has little effect on the activity of intact photosystem II, can reduce and inhibit the Mn cluster in both types of preparations. In contrast, a large reductant, hydroquinone, cannot access the majority of O(2)-evolving centers in citrate-treated preparations, while 23 and 17 kDa-depleted material is rapidly inactivated by the reductant. Incubation of the citrate-treated samples in high ( approximately 60 mM) concentrations of CaCl(2) restores 50% of the lost activity; this Ca(2+)-reconstituted activity is chelator-insensitive, indicating that rebinding of Ca(2+) restores the structural integrity of the O(2)-evolving complex. A characterization of Ca(2+) and Cl(-) affinities in steady-state activity assays shows that citrate-treated preparations exhibit a Cl(-) requirement similar to that of polypeptide-depleted photosystem II, while Ca(2+) reactivation of O(2) evolution appears to occur at two structurally distinct sites. One site exhibits a high Ca(2+) affinity, similar to that found in polypeptide-depleted samples, but a second, lower-affinity site also exists, with a K(M) that is approximately 10 times greater than that of the high-affinity site, which is associated with centers that retain the extrinsic polypeptides. These data indicate that citrate-induced Ca(2+) depletion causes release of the 23 and 17 kDa extrinsic polypeptides from some photosystem II reaction centers, and also modifies the structure of the polypeptide-retaining O(2)-evolving centers so that the Mn cluster is exposed to small, but not large, reductants. This change may be due to subtle modifications to the structure of the photosystem II extrinsic proteins that produces a new pathway between the solvent and the Mn cluster or, alternatively, to the opening of an existing channel in the intrinsic lumenal polypeptide domain, between the solvent and the Mn cluster, that is normally occluded by a bound Ca(2+) atom.
Vander Meulen
Kirk A
KA
Departments of Biology and Chemistry, University of Michigan, Ann Arbor, Michigan 48109-1048, USA.
Hobson
Amanda
A
Yocum
Charles F
CF
D002118:Calcium / Q000378:metabolism*; D002122:Calcium Chloride / Q000494:pharmacology; D019343:Citric Acid / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D010100:Oxygen / Q000378:metabolism*; D045322:Photosynthetic Reaction Center Complex Proteins / Q000302:isolation & purification / Q000378:metabolism*; D045332:Photosystem II Protein Complex; D012965:Sodium Chloride / Q000494:pharmacology; D018724:Spinacia oleracea / Q000378:metabolism
D045322:Photosynthetic Reaction Center Complex Proteins; D045332:Photosystem II Protein Complex; D019343:Citric Acid; D012965:Sodium Chloride; D002122:Calcium Chloride; D010100:Oxygen; D002118:Calcium