Construction and Characterization of Genetically Modified Synechocystis sp. PCC 6803 Photosystem II Core Complexes Containing Carotenoids with Shorter π-Conjugation than β-Carotene

Abstract

␤-Carotene has been identified as an intermediate in a secondary electron transfer pathway that oxidizes Chl Z and cytochrome b 559 in Photosystem II (PS II) when normal tyrosine oxidation is blocked. To test the redox function of carotenoids in this pathway, we replaced the -carotene desaturase gene (zds) or both the zds and phytoene desaturase (pds) genes of Synechocystis sp. PCC 6803 with the phytoene desaturase gene (crtI) of Rhodobacter capsulatus, producing carotenoids with shorter conjugated -electron systems and higher reduction potentials than ␤-carotene. The PS II core complexes of both mutant strains contain approximately the same number of chlorophylls and carotenoids as the wild type but have replaced ␤-carotene (11 double bonds), with neurosporene (9 conjugated double bonds) and ␤-zeacarotene (9 conjugated double bonds and 1 ␤-ionylidene ring). The presence of the ring appears necessary for PS II assembly. Visible and near-infrared spectroscopy were used to examine the light-induced formation of chlorophyll and carotenoid radical cations in the mutant PS II core complexes at temperatures from 20 to 160 K. At 20 K, a carotenoid cation radical is formed having an absorption maximum at 898 nm, an 85 nm blue shift relative to the ␤-carotene radical cation peak in the WT, and consistent with the formation of the cation radical of a carotenoid with 9 conjugated double bonds. The ratio of Chl ؉ /Car ؉ is higher in the mutant core complexes, consistent with the higher reduction potential for Car ؉ . As the temperature increases, other carotenoids become accessible to oxidation by P 680A carotenoid molecule is an extended polyene chain with a variety of end groups and has been thought of as a "molecular wire," because, in the one-electron oxidized form, or carotenoid radical cation, the hole is delocalized over the entire conjugated -system. Carotenoid radical cations have recently been observed in several photosynthetic pigmentprotein complexes, including PS II, 3 bacterial light-harvesting complexes LHC II, and recently in plant PsbS where they may have a role in non-photochemical energy quenching (1). Among photosynthetic reaction centers, carotenoid photooxidation is unique to PS II, which uses light energy to catalyze the oxidation of water to molecular oxygen. The process of water oxidation involves highly oxidizing species that can also oxidize a carotenoid molecule. Carotenoid radical cations have been identified as intermediates in the secondary electron transport pathway of PS II (2-6).Detergent-solubilized PS II core complexes contain up to 25 different integral membrane and extrinsic polypeptide subunits, including the D1/D2 complex and light-harvesting pigment-protein complexes CP43 (PsbC) and CP47 (PsbB). Light energy is absorbed by the chlorophyll-containing light-harvesting antenna and transferred to the primary donor chlorophyll(s) coordinated by the D1/D2 polypeptides. The term "PS II reaction centers" describes a structure that includes the D1 and D2 polypeptides, cytochrome b 559 (Cyt b ...

Isorenieratene Biosynthesis in Green Sulfur Bacteria Requires the Cooperative Actions of Two Carotenoid Cyclases

“…However, this level of enzymatic complexity is reminiscent of the complexity seen in converting phytoene to lycopene. Although a single enzyme, CrtI, can perform the same suite of reactions (22,25,34,46,50), two desaturases and at least one isomerase are required to convert phytoene into lycopene in GSB and most cyanobacteria (1,3,4,6,7,19,21,34,40,43).…”

“…These functions are very important in photosynthetic bacteria, and complete inactivation of the carotenogenic pathway in the cyanobacterium Synechocystis sp. strain PCC 6803 has been possible only in strains adapted to photoheterotrophic growth on glucose (3,4).…”

Characterization of the secondary electron-transfer pathway intermediates of photosystem II containing low-potential cytochrome b 559

“…species were identified (Car A at 982 nm and Car B at 1,027 nm). We note that for PS II containing carotenoids with less than 11 conjugated double bonds (Bautista et al 2005), there is no increase in absorption in the 1,030-1,100 nm region during extended illumination, which suggests that the absorption changes in this region observed here and in ferricyanide-treated PS II are due to the properties of PS II, and not due to the changes in the background absorption from water that could be produced by a heating artifact.…”

Carotenoids assist in assembly and functions of photosynthetic complexes in cyanobacteria

“…Moreover, genetic manipulation of the carotenoid synthesis suggested that PSII assembly requires the presence of a carotenoid with at least one β-ionylidene ring, which may play a structural role in early stages of assembly (Bautista et al 2005b). …”

“…Previously it has been suggested that β-carotene is vital for synthesis of D1 protein in the green alga, Chlamydomonas reinhardtii (Trebst and Depka 1997) as well as in the cyanobacterium, Synechocystis (Masamoto et al 2004). Moreover, genetic manipulation of the carotenoid synthesis suggested that PSII assembly requires the presence of a carotenoid with at least one β-ionylidene ring, which may play a structural role in early stages of assembly (Bautista et al 2005b). On the other hand, it is interesting to note that carotenoidless mutants of purple bacteria with type-2 RCs have normal photosynthesis.…”