THE ROLE OF REACTION-CENTER EXCITED-STATE EVOLUTION DURING CHARGE SEPARATION IN A RHODOBACTER-SPHAEROIDES MUTANT WITH AN INITIAL ELECTRON-DONOR MIDPOINT POTENTIAL 260 MV ABOVE WILD-TYPE

Femtosecond transient absorbance spectroscopy was performed on the tri ple hydrogen bond reaction center mutant [LH(L131) + LH(M160) + FH(M19 7)] of Rhodobacter sphaeroides which has a P/P+ midpoint potential 260 mV above wild type. The decay of the excited singlet state in this mu tant is kinetically complex with a dominant decay component of about 5 0 ps at 295 K. Charge separation to the state P(+)Q(A)(-) occurs with a quantum yield of 0.50 +/- 0.1 at 295 K and 0.10-0.15 at 20 K. The yi eld, rate of formation and spectra of states which are trapped when el ectron transfer to the quinone is blocked by quinone reduction compare d to the rate and yield of formation of P(+)Q(A)(-) in unreduced react ion centers suggest that evolution of the excited state is the rate li miting event in charge separation in triple mutant reaction centers. T he excited state that results from this evolution has spectral feature s which are remarkably similar to the initial excited singlet state fo und using R-26 reaction centers (R-26 reaction centers have essentiall y wild type photochemistry). The fact that the formation of this alter ed excited state is greatly slowed in a high P/P+ midpoint potential m utant suggests that the early excited state in wild type or R-26 react ion centers may have considerable P+ character. A consideration of the thermodynamics of the state P+BA- in this and related high potential mutants implies that a simple model in which P+BA- is formed as a disc rete electron transfer intermediate is not a viable description in the se mutants. Other factors such as reaction center heterogeneity or alt ernate electron transfer mechanisms must be invoked.