Abstract
The PsaC subunit of photosystem I (PS I) binds two [4Fe-4S] clusters, F A and F B, functioning as electron carriers between F X and soluble ferredoxin. To resolve the issue whether F A or F B is proximal to F X, we used single-turnover flashes to promote step-by-step electron transfer between electron carriers in control (both F A and F B present) and HgCl 2-treated (F B-less) PS I complexes from Synechococcus sp. PCC 6301 and analyzed the kinetics of P700 + reduction by monitoring the absorbance changes at 832 nm in the presence of a fast electron donor (phenazine methosulfate (PMS)). In control PS I complexes exogenously added ferredoxin, or flavodoxin could be photoreduced on each flash, thus allowing P700 + to be reduced from PMS. In F B-less complexes, both in the presence and in the absence of ferredoxin or flavodoxin, P700 + was reduced from PMS only on the first flash and was reduced from F X − on the following flashes, indicating lack of electron transfer to ferredoxin or flavodoxin. In the F B-less complexes, a normal level of P700 photooxidation was detected accompanied by a high yield of charge recombination between P700 + and F A − in the presence of a slow donor, 2,6-dichlorophenol-indophenol. This recombination remained the only pathway of F A − reoxidation in the presence of added ferredoxin, consistent with the lack of forward electron transfer. F A − could be reoxidized by methyl viologen in F B-less PS I complexes, although at a concentration two orders of magnitude higher than is required in wild-type PS I complexes, thus implying the presence of a diffusion barrier. The inhibition of electron transfer to ferredoxin and flavodoxin was completely reversed after reconstituting the F B cluster. Using rate versus distance estimates for electron transfer rates from F X to ferredoxin for two possible orientations of PsaC, we conclude that the kinetic data are best compatible with PsaC being oriented with F A as the cluster proximal to F X and F B as the distal cluster that donates electrons to ferredoxin.
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