Fluorescence spectra of native purple bacterial reaction center (RC) and bacterial pheophytin (Bphe) replaced RCs were obtained from 600 nm to 900 nm with a selective excitation at 597 nm. With the help of measuring the fluorescence from bacterial chlorophyll, bacterial pheophytin and plant pheophytin, the corresponding components in the RCs are classified for fluorescence emission. Results showed that pheophytin substitution influences the composition of fluorescence spectra. Therefore, four, three and two components were obtained from fluorescence spectra of native RC, Bphe B_replaced RC and Bphe A,B _replaced RC, respectively. Fluorescence components are well correlated to the binding of plant pheophytin. The decay of excited state of primary electron donor P in different RCs was also studied by measuring the fluorescence decay at 686.4, 674.1 and 681.1 nm, respectively. The decaying kinetics changed in different RCs, indicating that pheophytin replacement influenced the energy transduction and primary photochemical reaction in purple bacterial reaction centers.
Characterization of energy-transduction on die chloroplast thylakoid membranes from spinach (Spinacia oleracca L.) after thermal pretreatment was investigated. The related reactions of energy-transduction in chloroplasts were seriously affected by thermal pretreatment. The results were obtained as following: (1) The rate of cyclic photophosphorylation declined when the pretreatment temperature increased in the range of 25 to 45 degreesC. (2) The thermal pretreatment led to a decrease of the activity of thylakoid membrane-bounded ATPase. (3) Proton uptake of chloroplasts acid the fluorescence quenching of 9-aminoacridine (9-AA) in thylakoid membrane decreased after the thermal pretreatment, but addition of dicyclohexylcarbodiimide (DCCD) could partially restore the fluorescence quenching of 9-AA. (4) Both the rates of fast phase in electrochroism absorption change at 515 nm and the millisecond delayed light emission (ms-DLE) of chloroplast showed a progressive decrease upon raising the temperature of pretreatment. (5) Immunbloting analysis showed that the thermal pretreatment caused the changes of protein content and the electrophoresis mobility of thylakoid membrane-bound ATPase and its alpha -subunit. (6) If the temperature of pretreatment were higher than 33 degreesC, oxygen uptake of PS I -mediated in the samples was rapidly inhibited, but addition of sinapine into the reaction medium could partially restore the ability of oxygen uptake in the samples. These results are briefly discussed in relation to the change of permeability of thylakoid membranes, the dissociation of coupling factor complex as well as accumulation of the radicals in the thylakoid membranes after thermal pretreatment.
