Insertion of the N-terminal part of PsaF from Chlamydomonas reinhardtii into photosystem I from Synechococcus elongatus enables efficient binding of algal plastocyanin and cytochrome c(6)

A strain of the cyanobacterium Synechococcus elongatus was generated that e xpresses a hybrid version of the photosystem I subunit PsaF consisting of t he first 83 amino acids of PsaF from the green alga Chlamydomonas reinhardt ii fused to the C-terminal portion of PsaF from S, elongatus, The correspon ding modified gene was introduced into the genome of the psaF-deletion stra in FK2 by cointegration with an antibiotic resistance gene. The transforman ts express a new PsaF subunit similar in size to PsaF from C, reinhardtii t hat is assembled into photosystem I (PSI). Hybrid PSI complexes isolated fr om these strains show an increase by 2 or 3 orders of magnitude in the rate of P700(+) reduction by C, reinhardtii cytochrome c(6) or plastocyanin in 30% of the complexes as compared with wild type cyanobacterial PSI. The cor responding optimum second-order rate constants (k(2) = 4.0 and 1.7 x 10(7) M-1 S-1 for cytochrome c(6) and plastocyanin) are similar to those of PSI f rom C, reinhardtii. The remaining complexes are reduced at a slow rate simi lar to that observed with wild type PSI from S. elongatus and the algal don ors, At high concentrations of C, reinhardtii cytochrome c(6), a fast first -order kinetic component (t(1/2) = 4 mu s) is revealed, indicative of intra molecular electron transfer within a complex between the hybrid PSI and cyt ochrome c(6). This first-order phase is characteristic for P700(+) reductio n by cytochrome c(6) or plastocyanin in algae and higher plants. However, a similar fast phase is not detected for plastocyanin, Crosslinking studies show that, in contrast to PSI from wild type S, elongatus, the chimeric Psa F of PSI from the transformed strain cross-links to cytochrome c(6) or plas tocyanin with a similar efficiency as PsaF from C. reinhardtii PSI. Our dat a indicate that development of a eukaryotic type of reaction mechanism for binding and electron transfer between PSI and its electron donors required structural changes in both PSI and cytochrome c(6) or plastocyanin.