Daniel I. Arnon

Photochemical activity and components of membrane preparations from blue-green algae. I. Coexistence of two photosystems in relation to chlorophyll a and removal of phycocyanin.

Abstract Nostoc muscorum (Strain 7119) cells were disrupted and the accessory pigment phycocyanin was removed from membrane fragments by digitonin treatment. The phycocyanin-depleted membrane fragments retained both Photosystem I and Photosystem II activity, as evidenced by high rates of NADP+ photoreduction either by water or by reduced 2,6-dichlorophenolindophenol, indicating that phycocyanin is not an essential component for electron transport activity. No separation of the two photosystems was effected by the digitonin treatment. Even drastic digitonin treatments failed to diminish significantly the remarkably stable electron transport from water to NADP+. Action spectra and relative quantum efficiency measurements demonstrated the existence of both Photosystem I and Photosystem II in membrane fragments which contained chlorophyll a as the only significant light-absorbing pigment.

Photosynthesis by isolated chloroplasts. IV. General concept and comparison of three photochemical reactions.

Abstract 1. 1. Procedures are described for the preparation of chloroplasts capable of carrying out three photochemical reactions, each representing an increasingly complex phase of photosynthesis: photolysis of water (Hill reaction), esterification of inorganic phosphate into adenosine triphosphate (photosynthetic phosphorylation) and the reduction of carbon dioxide to the level of carbohydrates with a simultaneous evolution of oxygen. 2. 2. The three photochemical reactions were separable by variations in the technique for preparation of chloroplasts and by differential inhibition by several reagents. Inhibition of a more complex phase of photosynthesis does not affect the similar one which precedes it and, conversely, the inhibition of a simpler phase of photosynthesis is paralled by an inhibition of the more complex phase which follows. 3. 3. Reversible inhibition of CO 2 fixation and photosynthetic phosphorylation, but not of photolysis, by sulfhydryl group inhibitors suggests that sulfhydryl compounds (enzymes, cofactors, or both) are involved in phosphorylation and CO 2 fixation, but not in the primary conversion of light into chemical energy as measured by the Hill reaction. 4. 4. Evidence is presented in support of the conclusion that the synthesis of ATP by green cells occurs at two distinct sites: anaerobically in chloroplasts, by photosynthetic phosphorylation, and aerobically in smaller cytoplasmic particles, presumably mitochondria, by oxidative phosphorylation independent of light. 5. 5. A general scheme of photosynthesis by chloroplasts, consistent with these findings, is presented.

Oxidation-reduction potentials and stoichiometry of electron transfer in ferredoxins

The electron-carrier properties of a plant (Spinacia olarecea) and a bacterial (Clostridium pasteurianum) freedoxin were compared. The oxidation-reduction potentials, measured at different partial pressures of H2 and at different pH values, were found to be: E′0 = − 0.42 V for spinach ferredoxin and E′0 = − 0.39 V for clostridial ferredoxin. The oxidation-reduction potentials of spinach and clostridial ferredoxin were independent of pH in the ranges investigated: pH 6.67–8.19 for spinach ferredoxin and pH 6.13–7.41 for clostridial ferredoxin. The oxidation-reduction of clostridial ferredoxin, like that of spinach ferredoxin, was found to involve the transfer of 1 electron per molecule. These results are consistent with the view that, despite their diverse origin and function and their chemical differences, plant and bacterial ferredoxins have similar characteristics as electron carriers.

[39] Ferredoxins from photosynthetic bacteria, algae, and higher plants

Publisher Summary This chapter describes procedures for isolation of ferredoxin α from the photosynthetic green bacterium Chlorobium thiosulf atophilum ; ferredoxin α 1 from the photosynthetic purple sulfur bacterium Chromatium ; and ferredoxin β from higher plants (spinach and alfalfa leaves) and algae (blue-green, green, red, and brown types). In the course of ferredoxin assay, the method is based on the requirement of ferredoxin for the photochemical reduction of NADP by isolated spinach chloroplasts using reduced 2,6-dichlorophenolindophenol (DPIP) as the electron donor. The reaction is carried out in a cuvette (3-ml capacity, 1-cm light path) containing 2.4 ml of H 2 O and 0.1 ml each of buffer, DPIP, sodium ascorbate, chloroplasts, ferredoxin, and NADP. Because the reduction of ferredoxin by chloroplasts is light-dependent, strong illumination must be avoided during the preliminary steps of the assay. A method recently devised for isolation of ferredoxin from the photosynthetic purple nonsulfur bacterium, Rhodospirillum rubrum is described in the chapter.

Ferredoxin-dependent carbon assimilation in Rhodospirillum rubrum

SummaryEvidence has been presented that a soluble fraction from R. rubrum cells contains two new primary carboxylation reactions which depend on the reducing power of ferredoxin: (a) pyruvate synthase which brings about a synthesis of pyruvate from acetyl-CoA and CO2 and (b) α-ketoglutarate synthase which brings about a synthesis of α-ketoglutarate from succinyl-CoA and CO2. The soluble fraction of R. rubrum cells contains also a series of other enzymes which, together with the ferredoxin-dependent enzymes, constitutes a reductive carboxylic acid cycle—a new cyclic pathway for CO2 assimilation that was first found in the photosynthetic bacterium, Chlorobium thiosulfatophilum.

Photosynthesis by isolated chloroplasts V. Phosphorylation and carbon dioxide fixation by broken chloroplasts

Abstract 1. 1. Photosynthetic esterification of inorganic phosphate into adenosine triphosphate, and reduction of CO 2 to the level of carbohydrate, hitherto found to occur only in whole chloroplasts, have now been observed with chloroplasts broken by treatment with water. 2. 2. Broken chloroplasts retained only two of the three groups of enzymes contained in whole chloroplasts, namely, those controlling the photolysis of water and photosynthetic phosphorylation. At least some of the enzymes concerned in reduction of CO 2 were leached out by treating the chloroplasts with water, with the result that CO 2 fixation was completely abolished. 3. 3. On addition of the requisite cofactors, the capacity of broken chloroplasts for photosynthetic phosphorylation was the same as that of whole chloroplasts. 4. 4. The restoration to broken chloroplasts of the full capacity for photosynthetic CO 2 fixation of whole chloroplasts required the addition of pyridine nucleotides, adenosine triphosphate, and the soluble enzymes removed by water treatment of whole chloroplasts. 5. 5. An additional several-fold increase in the rate of CO 2 fixation by the reconstituted broken chloroplast system was obtained by the further addition of one of several compounds, principally phosphorylated sugars. This has resulted in a level of CO 2 fixation by broken chloroplasts which is much higher than that obtained with whole chloroplasts.

Photosynthesis by isolated chloroplasts: VII. Vitamin K and riboflavin phosphate as cofactors of cyclic photophosphorylation☆

Abstract Vitamin K substances and FMN appear to catalyze separate pathways of cyclic photophosphorylation. The FMN pathway shows a dependence on added TPN and greater sensitivity to inhibition by dinitrophenol and o -phenanthroline than the vitamin K pathway. Both pathways are inhibited by p -chloromercuribenzoate, gramicidin and methylene blue but not by arsenite or antimycin A. Cyclic photophosphorylation catalyzed by phenazine methosulfate resembled the vitamin K pathway in its independence from added TPN and resistance to inhibition by dinitrophenol and o -phenanthroline. The role of vitamin K in phosphorylations by plant and animal tissues is reviewed. A possible physiological role for cyclic photophosphorylation in photosynthesis of green plants is suggested.

Changes in glyceraldehyde phosphate dehydrogenase during the life cycle of a green plant

Abstract The pyridine nucleotide dependence of glyceraldehyde 3-phosphate dehydrogenase (GPD) was investigated throughout the life cycle of pea plants. The GPD of seeds and seedlings germinated in the dark was DPN-specific. In seedlings exposed to light, there appeared after the tenth day, a TPN-linked GPD. This new species of GPD was localized in the green shoot and was absent from roots. Seedlings germinated in the dark developed the TPN-linked GPD upon transfer to light. The light-induced TPN-linked GPD disappeared from the newly formed seed. In the life cycle of the plant from seed to seed, the GPD changes cyclically from obligate DPN dependence in the seed, to a multicomponent system in the shoot, functional with either DPN or TPN, and back to obligate DPN dependence in the newly formed seed.

Ferredoxins in light- and dark-grown photosynthetic cells with special reference to Rhodospirillum rubrum.

Two different ferredoxins (type I and type II) were isolated from photosynthetically grown Rhodospirillum rubrum cells. Type I had 6 non-heme irons and 6 acidlabile sulfides and an absorption ratio A385nm/A280nm = 0.72; type II had 2 non-heme irons and 2 acid-labile sulfides and an absorption ratio A385nm/A280nm = 0.52. The ferredoxins differed in amino acid composition and molecular weight (type I, 8700; type II, 7500). R. rubrum cells grown heterotrophically in the dark yielded type II ferredoxin; type I ferredoxin accompanied type II only when the cells were grown in the light. By comparison, spinach seedlings had only one type of ferredoxin whether germinated in the light or in the dark. R. rubrum appears to be the first organism from which two types of ferredoxin (one formed only in the light) were isolated. R. rubrum ferredoxin type II is the first instance of a ferredoxin from a photosynthetic bacterium that has a bacterial type absorption spectrum but a non-heme iron and labile sulfide content (two of each per mole) that is characteristic of plant ferredoxins.

On two photoreactions in System II of plant photosynthesis

Abstract Light-induced absorbance changes of cytochrome b559 and C550 in chloroplasts indicate that noncyclic electron transport from water to ferredoxin (Fd)-NADP+ is carried out solely by System II and includes not one but two photoreactions (IIa and IIb) that proceed effectively only in short-wavelength light. (C550 is a new chloroplast component identified by spectral evidence and distinct from cytochromes.) The evidence suggests that the two short-wavelength light reactions operate in series, being joined by a System II chain of electron carriers that includes (but is not limited to) C550, cytochrome b559, and plastocyanin (PC). H2O → IIbhv → C550 → cyt. b559 → PC → IIahv → Fd → NADP+ Photoreaction IIb involves an electron transfer from water to C550 that does not require plastocyanin and is the first known System II photoreaction resistant to inhibition by 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU) and o-phenanthroline. Cytochrome b559 is reduced by C550 in a reaction that is readily inhibited by DCMU or o-phenanthroline. Thus, the site of DCMU (and o-phenanthroline) inhibition of System II appears to lie between C550 and cytochrome b559. Photoreaction IIa involves an electron transfer from cytochrome b559 and plastocyanin to ferredoxin-NADP+.