The effect of far-red background-light. In chlorella or chloroplasts with added Hill - oxidants far-red actinic light between 700 and 730 mμ oxidizes Chl-aI; in the dark Chl-aI⊕ stays in this oxidized state. Actinic light in the range λ<700 mμ results also in an oxidation of Chl-aI, but in the dark Chl-aI⊕ is reduced. Therefore light of λ<700 mμ is channeled into two reaction centres. One part (hνI)is channeled to Chl-aI (with the effect of oxidation of Chl-aI), a second part (hνII) is channeled to a second reaction centre where it provides electrons for the reduction of Chl-aI⊕ [s. scheme (1)]. Under the influence of far-red background-light (700 -730 mμ) Chl-aI accumulates in its oxidized form. The magnitude of its oxidation depends on the intensity of the background-light. a) In soft far-red background-light practically no Chl-aI⊕ is accumulated (s. Fig. 1 a). A supplementary red flash (<700 mμ) at t1 with hνI and hνII-light should have the following effect: The hνI-light oxidizes Chl-aI immediately. The hνII-light provides electrons from a second reaction centre for the reduction of Chl-aI⊕. b) In stronger far-red background-light Chl-aI accumulates in its oxidized form (Fig. 1 b and Fig. 1 c). A supplementary red flash (<700 mμ) at t3 or t5 oxidizes the rest of Chl-aI immediately. Afterwards all Chl-aI⊕ is again reduced. - Changing from soft to stronger far-red background-light a shift from negative changes of absorption of Chl-aI to positive ones is aspected (Fig. 1 a′ and 1 b′). However, in chloroplasts without any added Hill- oxidants no shift takes place (Fig. 2 top). This indicates that no accumulation of Chl-aI⊕ has occurred. The reason is, that Chl-aI⊕ can be reduced by a backflow of electrons from the electron acceptor Z [s. scheme (2)]. Trapping the electrons of Z⊖ by electron acceptors (ox. SI) prevents the backflow [s. scheme (3)] and positive changes of absorption occur (Fig. 2 bottom). By shifting the changes of absorption of Chl-aI from negative to positive values it is possible to separate the difference-spectrum of Chl-ai from the overall difference-spectrum under complete natural conditions (s. Fig. 4 and 1. c. 1).Electron-acceptors of Z⊖. All substances surnamed under ox. SI (Table I and Fig. 11) trap electrons of Z⊖ (shift from negative to positive changes of absorption at 703 mμ). In this way it was shown, that also photosynthetic-pyridine-nucleotide-reductase (PPNR) - but not TPN alone - traps electrons from Ze. Therefore PPNR must contain a redox system [ferredoxin] [s. scheme (4)]. From the highest redox potential of ox. Si (methyl viologene) that of Z/Z⊖ (≲ -0,44 volt) was estimated.Water as the ultimate electron-donor for Chl-aI. CMU blocks oxidation of water. If water is the the ultimate electron donor for Chl-aI, turnover of Chl-aI should vanish in the presence of CMU [s. scheme (5)]. This is true for chlorella, but not for chloroplasts without ox. Si (Fig. 5 midth). The reason is again the backflow of electrons from Z⊖ to Chl-aI⊕ which keeps the cycle in action. Trapping the electrons of Z⊖ by addition of ox. Si results in the disappearence of the changes of Chl-ai (Fig. 5 bottom). In aged chloroplasts, where Chl-aI⊕ is supplied directly by electrons of PMSH⊖, addition of CMU should have no influence on the changes of Chl-aI (Fig. 6).a) Redox potential. By addition of different ratios of ferro/ferricyanide the ratio of Chl-aI/Chl-aI⊕ can be changed in the dark. This is shown by the dependence of the light induced changes at 703 mμ on the ratio of ferro/ferricyanide (Fig. 7). The oxidation of Chl-ai occurs by a single electron transfer. The redox potential of Chl-aI⊕/Chl-aI was determined to +0,46 ± 0,1 volt. b) The Chl-ai-reaction is stable between pʜ = 3 and pʜ = 11 (Fig. 8). c) The Chl-ai-reaction is stable up to 65°C (Fig. 9). d) Aging up to 7 days at 5°C has no influence upon the Chl-ai-reaction (Fig. 10).Reaction Scheme. The results are summarized in Fig. 11 and Table 2. In chlorella and fresh chloroplasts Chl-aI⊕ is reduced by electrons originating from water with the help of hνII-light. After suppressing water-oxidation (by aging, extraction of plastoquinone, treatment with digitonin, addition of CMU)1 Chl-aI⊕ can be reduced by backflow of electrons from Z⊖ or by added electron-donators as reduced PMS or reduced DPIP. The electrons of Z⊖ can be trapped by TPN (via ferredoxin) or substances surnamed under ox. SI.
Tuning the redox potential of the primary electron donor in bacterial reaction centers by manganese binding and light-induced structural changes
