Excited state energy transfer pathways in photosynthetic reaction centers. 3. Ultrafast emission from the monomeric bacteriochlorophylls

Ultrafast singlet excited state energy transfer occurs from the monomeric bacteriopheophytin (H) and bacteriochlorophyll (B) chromophores to the primary electron donor or special pair (P) in bacterial photosynthetic reaction centers. Because of rapid quenching of the singlet excited state of B by energy transfer to P, B-1 emission has not previously been observed in functional reaction centers. Using fluorescence upconversion, spontaneous fluorescence associated with the monomeric bacteriochlorophylls is observed for excitation of the monomeric bacteriochlorophylls and bacteriopheophytins at 85 K. The decay kinetics of the fluorescence match the kinetics of the rise of emission from P-1, the ultimate acceptor of singlet energy. Together with measurements of the time-resolved fluorescence anisotropy, the data suggest that B-1 is populated in the energy transfer pathway from H-1 to P. By exciting H in wild-type and in the reaction center mutant M182HL, where contributions from the chromophores in the B sites on the L and M sides are spectrally resolved, the amplitudes of the kinetic traces at several wavelengths between 790 and 825 nm can be used to construct the time-resolved emission spectrum of B-1. The Stokes shift of the accessory bacteriochlorophylls in wild-type on the femtosecond time scale is dose to zero, while for the monomeric bacteriopheophytin in the B-M binding site in the M182HL mutant, the Stokes shift is less than 100 cm(-1). These results have significant implications fur the mechanism of ultrafast energy transfer in the reaction center.