The D2 Protein in Photosystem II: Its Involvement in PS II Structure and Function

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This project has several objectives, all related to the function of D2, one of the photosystem II reaction center proteins in thylakoid membranes. Using site-directed mutagenesis and gene replacement in the D2-encoding pbsD1 gene from the cyanobacterium Synechocystis sp. PCC 6803, the first objective is to determine the role of residues near the C-terminus of D2 in the water-splitting process and in proper folding of D2. In addition, three- dimensional proximities of D2 residues involved in water splitting will be probed by analysis of spontaneous photoautotrophic mutants generated from obligate photoheterotrophic site-directed D2 mutants impaired in water splitting. Moreover, site-directed mutagenesis studies on electron transfer events at the acceptor side of photosystem II will emphasize experiments related to binding of additional chlorophyll(s) in the photosystem II reaction center complex as well as the role of the non-heme iron in photosystem II. Another objective, to be achieved in collaboration with Dr. Shestakov and his group in Moscow (USSR), is to identify other regions of the D2 protein that are important towards photosystem II function by random mutagenesis of psbD1. A successful completion of this project is anticipated to markedly increase the base of knowledge regarding the role of individual amino acid residues in function, folding and stable assembly of membrane proteins such as D2. Energy from the sun is captured for biological use by the process of photosynthesis. The process of light harvesting is carried out by two pigment-protein complexes, photosystems I and II. Light energy absorbed by photosystem I is used by the cell for carbon fixation and chemical energy production. Light energy absorbed by photosystem II is essential for regenerating photosystem I which loses electrons to cellular constituents. Photosystem II, in turn, is regenerated by the splitting of water, a process that liberates dioxygen and accounts for the presence of oxygen in the earth's atmosphere. This proposal describes molecular studies of a protein, D2, that is an essential part of photosystem II. These studies will help to elucidate how the process of water splitting occurs at the molecular level.//