Importance of the region including aspartates 57 and 60 of ferredoxin on the electron transfer complex with photosystem I in the cyanobacterium Synechocystis sp PCC 6803

Ferredoxin reduction by photosystem I has been studied by flash-absorption spectroscopy. Aspartate residues 20, 57, and 60 of ferredoxin were changed to alanine, cystein, arginine, or lysine. On the one hand, electron transfe r from photosystem I to all mutated ferredoxins still occurs on a microseco nd time scale, with halftimes of ferredoxin reduction mostly conserved comp ared to wild-type ferredoxin. On the other hand, the total amplitude of the fast first-order reduction varies largely when residues 57 or 60 are modif ied, in apparent relation to the charge modification (neutralized or invert ed). Substituting these two residues for lysine or arginine induce strong e ffects on ferredoxin binding (up to sixfold increase in K-D), whereas the s ame substitution on aspartate 20, a spacially related residue, results in m oderate effects (maximum twofold increase in K-D). In addition, double muta tions to arginine or lysine were performed on both aspartates 57 and 60. Th e mutated proteins have a 15- to 20-fold increased K-D and show strong modi fications in the amplitudes of the fast reduction kinetics. These results i ndicate that the acidic area of ferredoxin including aspartates 57 and 60, located opposite to the C-terminus, is crucial for high affinity interactio ns with photosystem I. (C) 2000 Academic Press.