STRUCTURAL MODEL OF CYTOCHROME B(559) IN PHOTOSYSTEM-II BASED ON A MUTANT WITH GENETICALLY FUSED SUBUNITS

Photosystem II is a reaction center protein complex located in photosy nthetic membranes of plants, algae, and cyanobacteria. Using light ene rgy, 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 b(559), a membrane-embedded heme protein with an unknown function. The cytochrome is unusual in t hat a heme links two separate polypeptide subunits, alpha and beta, ei ther as a heterodimer (alpha beta) or as two homodimers (alpha(2) and beta(2) and beta(2)) To determine the structural organization of cytoc hrome b(559) in the membrane, we used site-directed mutagenesis to fus e the coding regions of the two respective genes In the cyanobacterium Synechocystis sp. PCC 6803. In this construction, the C terminus of t he alpha subunit (9 kDa) is attached to the N terminus of the beta sub unit (5 kDa) to form a 14-kDa alpha beta fusion protein that is predic ted to have two membrane-spanning alpha-helices with antiparallel orie ntations. Cells containing the ap fusion protein grow photoautotrophic ally and assemble functional photosystem II complexes. Optical spectro scopy shows that the alpha beta fusion protein binds heme and is incor porated into photosystem II. These data support a structural model of cytochrome b(559) in which one heme is coordinated to an alpha(2) homo dimer and a second heme is coordinated to a beta(2) homodimer. In this model, each photosystem II complex contains two cytochrome b(559) hem es, with the alpha(2) heme located near the stromal side of the membra ne and the beta(2) heme located near the lumenal side.