Mutations in both sides of the photosystem I reaction center identify the phylloquinone observed by electron paramagnetic resonance spectroscopy

The core of photosystem I (PS1) is composed of the two related integral mem brane polypeptides, PsaA and PsaB, which bind two symmetrical branches of c ofactors, each consisting of two chlorophylls and a phylloquinone, that pot entially link the primary electron donor and the tertiary acceptor. In an e ffort to identify amino acid residues near the phylloquinone binding sites, all tryptophans and histidines that are conserved between PsaA and PsaB in the region of the 10th and 11th transmembrane alpha -helices were mutated in Chlamydomonas reinhardtii. The mutant PS1 reaction centers appear to ass emble normally and possess photochemical activity. An electron paramagnetic resonance (EPR) signal attributed to the phylloquinone anion radical(A(1)( -)) can be observed either transiently or after illumination of reaction ce nters with pre-reduced iron-sulfur clusters. Mutation of PsaA-Trp(693) to P he resulted in an inability to photo-accumulate A(1)(-), whereas mutation o f the analogous tryptophan in PsaB (PsaB-Trp(673)) did not produce this eff ect. The PsaA-W693F mutation also produced spectral changes in the time-res olved EPR spectrum of the P-700(+) A(1)(-) radical pair, whereas the analog ous mutation in PsaB had no observable effect. These observations indicate that the A(1)(-) phylloquinone radical observed by EPR occupies the phylloq uinone-binding site containing PsaA-Trp(693). However, mutation of either t ryptophan accelerated charge recombination from the terminal Fe-S clusters.