Conserved residue PsaB-Trp673 is essential for high-efficiency electron transfer between the phylloquinones and the iron-sulfur clusters in Photosystem I.

Despite the high level of symmetry between the PsaA and PsaB polypeptides in Photosystem I, some amino acids pairs are strikingly different, such as PsaA-Gly693 and PsaB-Trp673, which are located near a cluster of 11 water molecules between the A and A quinones and the F iron-sulfur cluster. In this work, we changed PsaB-Trp673 to PsaB-Phe673 in Synechocystis sp. PCC 6803. The variant contains ~ 85% of wild-type (WT) levels of Photosystem I but is unable to grow photoautotrophically. Both time-resolved and steady-state optical measurements show that in the PsaB-W673F variant less than 50% of the electrons reach the terminal iron-sulfur clusters F and F; the majority of the electrons recombine from A and A. However, in those reaction centers which pass electrons forward the transfer is heterogeneous: a minor population shows electron transfer rates from A and A to F slightly slower than that of the WT, whereas a major population shows forward electron transfer rates to F slowed to the ~ 10 µs time range. Competition between relatively similar forward and backward rates of electron transfer from the quinones to the F cluster account for the relatively low yield of long-lived charge separation in the PsaB-W673F variant. A higher water content and its increased mobility observed in MD simulations in the interquinone cavity of the PsaB-W673F variant shifts the pK of PsaB-Asp575 and allows its deprotonation in situ. The heterogeneity found may be rooted in protonation state of PsaB-Asp575, which controls whether electron transfer can proceed beyond the phylloquinone cofactors.