The oxidation state of the photosystem II manganese cluster influences thestructure of manganese stabilizing protein

Exposure of photosystem II membranes to trypsin that has been treated to in hibit chymotrypsin activity produces limited hydrolysis of manganese stabil izing protein. Exposure to chymotrypsin under the same conditions yields su bstantial digestion of the protein. Further probing of the unusual insensit ivity of manganese stabilizing protein to trypsin hydrolysis reveals that i ncreasing the temperature from 4 to 25 degreesC will cause some acceleratio n in the rate of proteolysis. However, addition of low (100 muM) concentrat ions of NH2OH, that are sufficient to reduce, but not destroy, the photosys tem II Mn cluster, causes a change in PS II-bound manganese stabilizing pro tein that causes it to be rapidly digested by trypsin. Immunoblot analyses with polyclonal antibodies directed against the N-terminus of the protein, or against the entire sequence show that trypsin cleavage produces two dist inct peptide fragments estimated to be in the 17-20 kDa range, consistent w ith proposals that there are 2 mol of the protein/mol photosystem II. The c orrelation of trypsin sensitivity with Mn redox state(s) in photosystem II suggest that manganese stabilizing protein may interact either directly wit h Mn, or alternatively, that the polypeptide is bound to another protein of the photosystem II reaction center that is intimately involved in binding and redox activity of Mn. (C) 2001 Elsevier Science B.V. All rights reserve d.