Deletions in the lumenal domains of the D1 protein that prevent the assembly of photosystem II in Synechocystis sp. PCC 6803

Abstract

The photosystem II (PSII) proteins, D1 and CP47, contribute hydrophilic domains to the water-oxidizing complex (WOC). We have constructed a deletion mutant system that allows us to introduce mutations into psbA2, encoding D1. Initially we introduced short deletions of 3 to 6 amino acids that targeted alkoxo and phenoxo groups from Ser and Tyr residues creating the strains: D1:?(Y73-S79); D1:?(V82-S86); D1:?(Y107-G109); D1:?(L174-S177) and D1:?(F302-S305); however, PSII was not assembled in these mutants. In addition, we created the mutant D1:T292L which exhibited a phenotype similar to wild type. However, removal of PSII-V (or cytochrome c-550) abolished photoautotrophic growth in the D1:T292L:?PSII-V strain in a pH dependent fashion: no growth was observed at pH 8.0 but it was restored at pH 10. The Arg-384 to Val-392 region of CP47 has been shown to influence the tight binding of PSII-O (or the manganese-stabilizing protein) to the WOC. Mutations were introduced that created the strains: CP47:E387R; CP47:S388A; CP47:K389E; CP47:K389Q and CP47:S391R. Each of these mutants were characterized in the presence or absence of PSII-O, PSII-U and PSII-V; and, mutations at Ser-388, Lys-389 and Ser-391 resulted in a phenotype similar to wild type or a control strain lacking the corresponding extrinsic protein. However, deletion of PSII-V from CP47:E387R resulted in an obligate photohetrotrophic strain that did not assemble PSII when grown at pH 8.0. However, growth of CP47:E387R:?PSII-V cells at pH 10 restored PSII assembly and photoautotrophic growth. We are testing if mutations at D1:Thr-292 and CP47:Glu-387 are additive when introduced in the same strain.