Abstract
Manganese (Mn) and calcium (Ca) are both metal cofactors required for photosynthetic oxygen evolution. The functional roles for these ions in the O2-evolving reactions are not completely known. They are studied by comparative spectroscopic measurements between intact and metal-depleted samples. In this chapter, we outline three experimental procedures used for the various removal of Mn and Ca from photosystem (PS) II-containing (i.e,. O2-evolving) preparations: the complete Mn extraction using a strong alkaline Ches buffer (pH 9.4)/MgCl2 wash, partial Mn extraction using a mild hydroxylamine (pH 6.8) wash, and specific Ca extraction through a low pH/citrate (pH 3) wash. The O2 evolution activities (measured by a Clarke-type oxygen electrode), protein composition (determined by sodium dodecyl sulfate- polyacrylamide gel electrophoresis), and the relative Mn and Ca content (measured by atomic absorption spectroscopy) are reported for each extraction procedure.
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