Abstract
Photosystem I functions as a light-driven plastocyanin-ferredoxin oxidoreductase in the photosynthetic membranes of cyanobacteria and chloroplasts. A mutant strain of the cyanobacterium Synechocystis sp. PCC 6803 that contains a deletion of the psaF gene and a transcriptionally inactive psaJ gene has assembled photosystem I complexes that lack PsaF, a lumenal protein and PsaJ, a 4-kDa hydrophobic protein. The cells of the mutant and wild type strains have similar rates of photosynthetic electron transfer and P700+ rereduction under linear and cyclic electron transfer conditions. Analysis of flash-induced absorption transients at 700 nm demonstrate that the absence of PsaF in purified mutant photosystem I did not affect the rate of P700 rereduction by cytochrome c553. Therefore, PsaF is not essential for docking of cytochrome c553. We also studied the organization of the proteins of mutant and wild type photosystem I by comparing their accessibility to digestion by thermolysin or to removal by 1 M NaI. The PsaA-PsaB subunits were more easily degraded by thermolysin in the mutant photosystem I. Thermolysin cleavage of PsaB yielded two major fragments that were immunoreactive with an antibody raised against the C terminus of PsaB. The N termini of these PsaB peptides mapped at Ile482 and Ile498 residues, thus identifying a surface-exposed domain of the core of photosystem I. The PsaE subunit could be removed by 1 M NaI and was rapidly digested by thermolysin in the mutant but not in the wild type photosystem I. Therefore, PsaF and PsaJ subunits of photosystem I have dispensable accessory roles in the function and organization of the complex.
Highlights
Analysis of flash-induced absorption transients at 700 n m demonstrate that the absence of PsaF’ in purified mutant photosystem I did not affect the rate of P700 rereduction by cytochrome csm
We studied cyclic electron flow since cyanobacterial mutants lacking PsaE show impaired cyclic electron flow [11].The PsaL subunitdoes not participatein the photochemical or electron transportprocesses [12] but is required for the formation of PS I trimers
We compared photosynthetic electron transfer rates and P700 rereduction kineticswihnole cells and purified PS I complexes of wild typeand AFK6
Summary
Containing 10pg/ml chlorophyll-containingPS I, 2 m~ ascorbate, and 1 m~ methyl viologen. Advances in resolution of low mass polypeptides have identified three new subunits, through Centricon-30 and Centricon-10, and extensively washed with PsaJ, PsaI, and PsaM, in PS I of Synechocystis sp. Isolation of the Photosynthetic Membranes and Purification of PS I-Synechocystis sp. PCC 6803 cultures of both the wild type and mutant strains were harvested and resuspended in 0.4 M sucrose, 10 m~ NaC1,200 p~ phenylmethylsulfonyl fluoride, 5 m~ benzamidine, and 10. P m diameter) was added, and the cells werebroken with a bead beater (Biospec)after several breaking-cooling cycles. Thylakoid membranes were separated from unbroken cells, washed extensively with, and resuspended in 0.4 M sucrose, 10 m~ NaCl, and 10 m~ MOPS, pH 7.0.To isolate PS I, the thylakoid membranes were solubilized with Triton. PCC 6803 codes for a 4442-Da polypeptide of 40 amino acids. X-100, followed by DEAF,-cellulosechromatography and sucrose gradi- tein of Synechocystisis remarkably conservedwhen compared
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