STRAINS OF SYNECHOCYSTIS SP PCC-6803 WITH ALTERED PSAC .2. EPR AND OPTICAL SPECTROSCOPIC PROPERTIES OF F-A AND F-B IN ASPARTATE, SERINE, AND ALANINE REPLACEMENTS OF CYSTEINE-14 AND CYSTEINE-51

A psaC deletion mutant of the unicellular cyanobacterium Synechocystis sp, PCC 6803 was utilized to incorporate site-specific amino acid sub stitutions in the cysteine residues that ligate the F-A and F-B iron-s ulfur clusters in Photosystem I (PS I). Cysteines 14 and 51 of PsaC we re changed to aspartic acid (C14D(PsaC), C51D(PsaC), C14D/C51D(PsaC)), serine (C14S(PsaC), C51S(PsaC)), and alanine (C14A(PsaC), C51A(PsaC)) , and the properties of F-A and F-B were characterized by electron par amagnetic resonance spectroscopy and time-reserved optical spectroscop y, The C14D(PsaC)-PS I and C14S(PsaC)-PS I complexes showed high level s of photoreduction of F-A with g values of 2.045, 1.944, and 1.852 af ter illumination at 15 K, but there was no evidence of reduced F-B in the g = 2 region, The C51D(PsaC)-PS I and C51S(PsaC)-PS I complexes sh owed low levels of photoreduction of F-B with g values of 2.067, 1.931 , and 1.881 after illumination at 15 K, but there was no evidence of r educed F-A in the g = 2 region, The presence of F-B was inferred in C1 4D(PsaC)-PS I and C14S(PsaC)-PS I, and the presence of F-A was inferre d in C51D(PsaC)-PS I and C51S(PsaC)-PS I by magnetic interaction in th e photoaccumulated spectra and by the equal spin concentration of the irreversible P700(+) cation generated by illumination at 77 K, Flash-i nduced optical absorbance changes at 298 K in the presence of a fast e lectron donor indicate that two electron accepters function after F-X in the four mutant PS I complexes at room temperature, These data sugg est that a mixed-ligand [4Fe-4S] cluster is present in the mutant site s of CI LX-PS I and C5LX-PS I (where X = D or S), but that the propose d spin state of S = 3/2 renders the resonances undetectable in the g 2 region, The C14A(PsaC)-PS I, C51A(PsaC)-PS I and C14D/C51D(PsaC)-PS I complexes show only the photoreduction of F-X, consistent with the ab sence of PsaC. These results show that only those PsaC proteins that c ontain two [4Fe-4S] clusters are capable of assembling onto PS I cores in vivo.