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Structural model of cytochrome b559 in photosystem II based on a mutant with genetically fused subunits
- McNamara, V.P., Sutterwala, F.S., Pakrasi, H.B., Whitmarsh, J.
- Proceedings of the National Academy of Sciences of the United States of America 1997 v.94 no.25 pp. 14173-14178
- structural genes, Synechocystis, heme, recombinant proteins, thylakoids, site-directed mutagenesis, photosystem II, mutants, cytochrome b, chemical structure, immunocytochemistry
- Photosystem II is a reaction center protein complex located in photosynthetic membranes of plants, algae, and cyanobacteria. Using light energy, photosystem II catalyzes the oxidation of water and the reduction of plastoquinone, resulting in the release of molecular oxygen. A key component of photosystem II is cytochrome b559, a membrane-embedded heme protein with an unknown function. The cytochrome is unusual in that a heme links two separate polypeptide subunits, alpha and beta, either as a heterodimer (alpha beta) or as two homodimers (alpha 2 and beta 2). To determine the structural organization of cytochrome b559 in the membrane, we used site-directed mutagenesis to fuse the coding regions of the two respective genes in the cyanobacterium Synechocystis sp. PCC 6803. In this construction, the C terminus of the alpha subunit (9 kDa) is attached to the N terminus of the beta subunit (5 kDa) to form a 14-kDa alpha beta fusion protein that is predicted to have two membrane-spanning alpha-helices with antiparallel orientations. Cells containing the alpha beta fusion protein grow photoautotrophically and assemble functional photosystem II complexes. Optical spectroscopy shows that the alpha beta fusion protein binds heme and is incorporated into photosystem II. These data support a structural model of cytochrome b559 in which one heme is coordinated to an alpha 2 homodimer and a second heme is coordinated to a beta 2 homodimer. In this model, each photosystem II complex contains two cytochrome b559 hemes, with the alpha 2 heme located near the stromal side of the membrane and the beta 2 heme located near the lumenal side.