AbstractWe investigated the influence of a range of factors—temperature, redox midpoint potential of an electron carrier, and protein dynamics—on nanosecond electron transfer within a protein. The model reaction was back electron transfer from a bacteriopheophytin anion, H_A⁻, to an oxidized primary electron donor, P⁺, in a wild type Rhodobacter sphaeroides reaction center (RC) with a permanently reduced secondary electron acceptor (quinone, Q_A⁻). Also used were two modified RCs with single amino acid mutations near the monomeric bacteriochlorophyll, B_A, located between P and H_A. Both mutant RCs showed significant slowing down of this back electron transfer reaction with decreasing temperature, similar to that observed with the wild type RC, but contrasting with a number of single point mutant RCs studied previously. The observed similarities and differences are explained in the framework of a (P⁺B_A⁻ ↔ P⁺H_A⁻) equilibrium model with an important role played by protein relaxation. The major cause of the observed temperature dependence, both in the wild type RC and in the mutant proteins, is a limitation in access to the thermally activated pathway of charge recombination via the state P⁺B_A⁻ at low temperatures. The data indicate that in all RCs both charge recombination pathways, the thermally activated one and a direct one without involvement of the P⁺B_A⁻ state, are controlled by the protein dynamics. It is concluded that the modifications of the protein environment affect the overall back electron transfer kinetics primarily by changing the redox potential of B_A and not by changing the protein relaxation dynamics.