Summary
The light-induced formation of NADH by whole cells of Rhodopseudomonas spheroides has been followed fluorimetrically and found to lag slightly behind cytochrome c oxidation. The uncoupler, FCCP1, abolished NADH formation which was also inhibited by HOQNO1. Electron flow from NADH to oxygen or cytochrome c was inhibited in chromatophores of R. spheroides by HOQNO, antimycin A and rotenone. From the known properties of the inhibitors used it is deduced that NADH formation in the light is dependent upon reversed electron flow. No light-induced formation of NAD(P)H by whole cells or chromatophores of Chlorobium thiosulfatophilum was detected either fluorimetrically or by extraction followed by enzymic assay although cytochrome c oxidation was extensive in whole cells. Extracts of C. thiosulfatophilum catalysed the rapid reduction of endogenous or mammalian cytochrome c; unlike R. spheroides this activity was found almost entirely in the soluble fraction and was insensitive to HOQNO, antimycin A and rotenone. No cytochrome b was detected in C. thiosulfatophilum by difference spectroscopy of pyridine haemochromes of acetone powders. The K m for NADH of NADH-cytochrome c reductase in both organisms was about 3 μmol; the reductase was inhibited by NAD. The rates of NADPH-cytochrome c reductase in R. spheroides particles were too low for K m determination; for C. thiosulfatophilum particles the K m for NADPH was about 300 μmol. The addition of NADH to soluble extracts of either organism caused the reduction of endogenous flavin that was reoxidised by ferricyanide. The NADH-cytochrome c reductase of C. thiosulfatophilum was not separated from ferredoxin on a DEAE column. It is concluded that in C. thiosulfatophilum the formation of NADH in an energy-linked reaction is unlikely; the possibility of a cyclic electron flow involving chlorophyll, ferredoxin, flavoprotein and cytochrome c is discussed.
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References
Buchanan, R. B., Matsubara, H., Evans, M. C. W.: Ferredoxin from the photosynthetic bacterium Chlorobium thiosulfatophilum—a link to ferredoxins from non photosynthetic bacteria. Biochim. biophys. Acta (Amst.) 189, 46–53 (1969).
Cartier, P. H.: Dosage des pyridine nucléotides oxydés et reduits dans le sang et les tissues animaux. Europ. J. Biochem. 4, 247–255 (1968).
Chance, B.: Rapid and sensitive spectrophotometry—A double beam apparatus. Rev. Sci. Instr. 22, 634–638 (1951).
Evans, M. C. W., Buchanan, R. B.: Photoreduction of ferredoxin and its use in carbon dioxide fixation by a sub-cellular system from a photosynthetic bacterium. Proc. nat. Acad. Sci. (Wash.) 53, 1420–1425 (1965).
——, Arnon, D. I.: A new ferredoxin—dependent carbon reduction cycle in a photosynthetic bacterium. Proc. nat. Acad. Sci. (Wash.) 55, 928–934 (1966).
Estabrook, R. W., Williamson, J. R., Frenkel, R., Maitra, P. K.: The fluorimetric determination of mitochondrial adenine and pyridine nucleotides. In: Methods in Enzymology, Vol. X, pp. 474–482. Eds. R. W. Estabrook and M. E. Pullman. New York-London: Academic Press 1967.
Forti, G.: Studies on NADPH-cytochrome f reductase of chloroplasts. In: Energy conservation by the photosynthetic apparatus. Brookhaven Symposia in Biology No. 19, pp. 195–201 (1966).
Foust, G. P., Mayhew, S. G., Massey, V.: Complex formation between ferredoxin-NADP reductase and electron transfer proteins. J. biol. Chem. 244, 964–970 (1969).
Garland, P. B., Clegg, R. A., Light, P. A., Ragan, C. I.: Mechanisms and inhibitors acting on electron transport and energy conservation between NADH and the cytochrome chain. In: Inhibitors: Tools in Cell Research, 20th Mosbach Colloquium, pp. 217–246. Eds. Th. Bucher and H. Sies. Berlin-Heidelberg-New York: Springer 1969.
Horio, T., Bartsch, R. G., Kakuno, T., Kamen, M. D.: Two NADH dehydrogenases from the photosynthetic bacterium Rhodospirillum rubrum. J. biol. Chem. 244, 5899–5909 (1969).
Jackson, J. B., Crofts, A. R.: Energy-linked reduction of NAD(P) in cells of Rhodospirillum rubrum. Biochem. biophys. Res. Commun. 32, 908–915 (1968).
Jones, O. T. G.: Multiple light-induced reactions of cytochromes b and c in Rhodopseudomonas spheroides. Biochem. J. 114, 793–799 (1969).
Keister, D. L., Yike, N. J.: Energy linked reactions in photosynthetic bacteria: succinate-linked ATP-driven NAD reduction by Rhodospirillum rubrum chromatophores. Arch. Biochem. 121, 415–422 (1967).
Klemme, J.-H.: Studies on the mechanism of NAD-photoreduction by chromatophores of the facultative photo-heterotroph Rhodopseudomonas capsulata. Z. Naturforsch. 24b, 67–76 (1969).
Kröger, A., Klingenberg, M.: On the role of ubiquinone in mitochondria. II. Redox reactions of ubiquinone under the control of oxidative phosphorylation. Biochem. Z. 344, 317–336 (1966).
Nishimura, M.: The effects of HOQNO and antimycin A on the photosynthetic and respiratory electron-transfer systems in photosynthetic bacteria. Biochim. biophys. Acta (Amst.) 66, 17–20 (1963).
Sabo, D. J., Orlando, J. A.: Isolation, purification and some properties of NADPH-cytochrome c2 reductase from Rhodopseudomonas spheroides. J. biol. Chem. 243, 3742–3749 (1968).
Tagawa, K., Arnon, D. I.: Ferredoxins as electron carriers in photosynthesis. Nature (Lond.) 195, 537–543 (1962).
Thore, A., Keister, D. L., San Pietro, A.: Studies on the respiratory system of aerobically (dark) and anaerobically (light) grown Rhodospirillum rubrum. Arch. Mikrobiol. 67, 378–396 (1969).
Trebst, A., Burba, M.: Disalicylidene propane diamine inhibition of photosynthesis in isolated chloroplasts and Chlorella. Z. Pflanzenphysiol. 57, 419–430 (1967).
Worcel, A., Goldman, D. S., Cleland, W. W.: An allosteric reduced NAD oxidase from Mycobacterium tuberculosis. J. biol. Chem. 240, 3399–3407 (1965).
Yamanaka, T., Kamen, M. D.: A NADP reductase, a NADH-dye reductase, and a non-haem iron protein isolated from a faculative photoheterotroph Rhodopseudomonas palustris. Biochim. biophys. Acta (Amst.) 131, 317–329 (1967).
Yamashita, J., Kamen, M. D., Horio, T.: Effect of oligomycin on NADH oxidation and its coupled phosphorylation with the particulate fraction from dark aerobically grown Rhodospirillum rubrum. Arch. Mikrobiol. 66, 304–314 (1969).
Yang, C.-C., Legallais, V.: A rapid and sensitive recording spectrophotometer for the visible and ultraviolet region. Rev. Sci. Instr. 25, 801–813 (1954).
Yocum, C. F., San Pietro, A.: Ferredoxin reducing substance (FRS) from spinach. Biochem. biophys. Res. Commun. 36, 614–620 (1969).
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Jones, O.T.G., Whale, F.R. The oxidation and reduction of pyridine nucleotides by Rhodopseudomonas spheroides and Chlorobium thiosulfatophilum . Archiv. Mikrobiol. 72, 48–59 (1970). https://doi.org/10.1007/BF00411014
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DOI: https://doi.org/10.1007/BF00411014