MUTATION VAL235ALA WEAKENS BINDING OF THE 33-KDA MANGANESE-STABILIZING PROTEIN OF PHOTOSYSTEM-II TO ONE OF 2 SITES

The 33-kDa protein of the photosynthetic O-2-evolving complex, also kn own as manganese stabilizing protein, contributes to the structural st ability of the photosystem II tetranuclear Mn cluster and stimulates t he water-oxidizing activity of this cluster. Quantification of extrins ic polypeptides in photosystem II has yielded data that support stoich iometries of either one or two copies of each protein per photosystem II reaction center. We recently described the cold-sensitive assembly of a mutant 33-kDa protein with a single amino acid replacement (Val23 5Ala) [Betts, S. D., Ross, J. R., Pichersky, E., & Yocum, C. F. (1996) Biochemistry 35, 6302-6307]. We have extended the characterization of this mutation. When photosystem II membranes depleted of the 33 kDa e xtrinsic protein are exposed to mixtures of wild type and Val235Ala ma nganese stabilizing protein, binding of the wild type protein is stron gly preferred. If, however, protein containing the Val235Ala mutation is first bound to photosystem II only half of this protein (about 1 mo l/mol of photosystem II reaction centers) is susceptible to displaceme nt by the wild type protein, even after multiple exposures to the latt er. These results support the conclusion that 2 mol of manganese stabi lizing protein are bound per reaction center. Our data show as well th at the mutant 33-kDa protein competes with the wild type protein for a t least one of two binding sites on photosystem II and that the mutant protein binds tightly to only one of two sites. These results demonst rate that the two binding sites on photosystem II for the 33-kDa prote in have different properties with respect to recognition and binding o f this protein.