Electron transfer towards the RCI-type photosystem in the green sulphur bacterium Chlorobium limicola forma thiosulphatophilum studied by time-resolved optical spectroscopy in vivo.
Flash-induced spectral changes in the wavelength region of the alpha-peaks of heme proteins and in the time domain from microseconds to seconds have been recorded on whole cells of the green sulphur bacterium Chlorobium limicola forma thiosulfatophilum. Extensive flash-excitation by trains of flashes resulted in oxidation of 7-8 c-type heme molecules/photosynthetic reaction centre. The complement of heme species was found to be spectrally heterogeneous allowing the study of electron transfer events induced by an isolated single-turnover flash. Under single-flash conditions, a c553 heme was seen to become oxidised with tau = 30 micros, concommitant with the reduction of the primary donor of the reaction centre. Subsequently, the alpha-peak of the photooxidised heme broadened and shifted towards longer wavelengths (tau = 70 micros) indicating equilibration of the positive charge over two differing heme species. In the time domain t > 1 ms, rereduction of c-type hemes was seen to be paralleled by a blue shift and further broadening of the alpha-peak. Concommitantly, b-type hemes were observed to first become reduced (within a few milliseconds), then over-oxidised (t > 200 ms) and eventually rereduced to their redox state prior to the flash. The results obtained are discussed with respect to the question of the identity of the immediate electron donor to the photosynthetic reaction centre and with respect to the involvement of a cytochrome bc complex in photo-induced electron transport of green sulphur bacteria.