| The purple photosynthetic bacterium Rb. sphaeroides is used to investigate processes of photosynthesis. Electric fields are applied along the membrane normal of Rb. sphaeroides chromatophore vesicles by the creation of a potassium chloride gradient across the membrane. The fluorescence emission is monitored as a function of the applied electric field potential.;The electric-field effect on the fluorescence emission is maximum (approximately 15% per 100 mV) in the Fm state, when the reaction center special pair is oxidized (P+) and the reaction center cannot perform charge separation. The fluorescence changes we observe under these conditions are consistent with an electric-field effect directly on the fluorescence emission of the LH1 and LH2 antenna complexes. This theory is supported by an electric-field effect on the fluorescence emission of LM1.1, a mutant Rb. sphaeroides strain which expresses LH1 and LH2 but not reaction centers. In the wild-type Fo state, when the reaction center is capable of charge separation, the electric-field effect is diminished (approximately 8% per 100 mV). Assuming that the electric-field effect on the reaction center is separate from the electric-field effect on the antenna complexes, subtraction of electric-field effect on the Fm state from the electric-field effect on the Fo state yields an estimate of the electric-field effect on the reaction center. Comparison of this effect with previous theoretical work supports the superexchange mechanism of charge separation in the reaction center.;Excitation equilibration is investigated in several mutant strains of Rb. sphaeroides which lack reaction centers, as well as the wild type. The Kennard-Stepanov analysis of absorption and fluorescence spectra is applied to membrane preparations of the mutants RC-IA, BALM/LH2, and LM1.1, which express LH1, LH2, and both LH1 and LH2, respectively. Good agreement with the Kennard-Stepanov (KS) relation is found over the majority of the spectral range, indicating rapid and complete excitation equilibration prior to fluorescence emission. One exception is a deviation on the red edge of the spectra, indicating a possible "dark" state in that spectral region which absorbs but does not fluoresce with the expected intensity.;The wild-type Rb. sphaeroides membrane preparation also shows a red-edge anomaly in the KS spectral analysis. Additionally, there is a second anomaly in the central spectral region which is not seen for mutants lacking a reaction center. Therefore we conclude that the presence of a reaction center disrupts the complete equilibration of excitation prior to fluorescence emission. Notably, this disruption of excitation equilibration is not observed in any previous KS analysis of PSI and PSII. |
