The role of cyclic photophosphorylation in vivo

When cyclic photophosphorylation is inhibited in Chlorella vulgaris cells by carbonylcyanide-trifluoromethoxy phenylhy-drazone, photosynthetic CO₂-fixation under anaerobic conditions exhibits a distinct lag. Under the same conditions, the light-dependent formation of ribulose diphosphate shows also this lag. It is concluded that cyclic photophosphorylation is required to fill up the pools of phosphorylated intermediates of the Calvin cycle at a time when noncyclic photophosphorylation cannot yet efficiently operate. Under aerobic conditions, the initial energy demand can be accommodated by respiratory ATP or cyclic photophosphorylation or both. Evidence for stoichiometric participation of cyclic photophosphorylation in photosynthesis is still lacking.     

Photosynthesis, Ribulose-1,5-bisphosphate Carboxylase, Electron Transport, and Ribulose 1,5-Bisphosphate of Virescent and Normal Green Wheat Leaves

CO₂ gas exchange, ribulose-1,5-bisphosphate, and electron transport have been measured in leaves of a yellow-green mutant of wheat (Triticum durum var Cappelli) and its wild type strain grown in the field. All these parameters, expressed on leaf area basis, were similar in both genotypes except electron transport which was more than double in the wild type. These results, treated according to a recent photosynthesis model for C₃ plants, seem to indicate that the electron transport rate of mutant leaves is not sufficient to support the carboxylation derived through both the assimilation rate and the in vitro ribulose-1,5-bisphosphate carboxylase activity. It is suggested that under our experimental conditions photosynthetic electron transport is not the sole energy-dependent determinant of ribulose-1,5-bisphosphate regeneration in the mutant.     

A Mutant of Arabidopsis thaliana Which Lacks Activation of RuBP Carboxylase In Vivo

A mutant of Arabidopsis thaliana has been isolated in which ribulose-1,5-bisphosphate carboxylase is present in a nonactivatable form in vivo. The mutation appears to affect carboxylase activation specifically, and not any other enzyme of the photosynthesis or photorespiratory cycles. The effect of the mutation on carboxylase activation is indirect, inasmuch as the properties of ribulose-1,5-bisphosphate carboxylase purified from the mutant are not distinguishable from those of the wild type enzyme. The mutant requires high levels of atmospheric CO₂ for growth because photosynthesis is severely impaired in atmospheres containing normal levels of CO₂, irrespective of the atmospheric O₂ concentration. In this respect, the mutant is distinguished from previously described high-CO₂ requiring mutants of Arabidopsis which have defects in photorespiratory carbon or nitrogen metabolism.     

The role of photophosphorylation in SO2 and SO 3 2- inhibition of photosynthesis in isolated chloroplasts

Sulphur dioxide inhibits noncyclic photophosphorylation in isolated envelope-free chloroplasts. This inhibition was shown to be reversible and competitive with phosphate, with an inhibitor constant of K_i=0.8mM. The same inhibition characteristics were observed when phosphoglycerate (PGA)- or ribulose-1,5-bisphosphate (RuBP)-dependent oxygen evolution was examined in a reconstituted chloroplast system in the presence of SO ₃ ^2- . Using an ATP-regenerating system (phosphocreatine-creatine kinase), it was demonstrated that the inhibition of PGA-dependent oxygen evolution is solely the result of inhibited photophosphorylation. It is concluded that at low SO₂ and SO ₃ ^2- concentrations the inhibition of photophosphorylation is responsible for the inhibition of photosynthetic oxygen evolution.Abbreviations Chl chlorophyll - PGA D-3-phosphoglyceric acid trisodium salt - Pi inorganic phosphate - RuBP D-ribulose-1,5-bisphosphoric acid tetrasodium salt     

Comparison of Properties of the Proteolytic Degradation of Unassembled Nuclear-encoded Subunits of Ribulose-1,5-bisphosphate Carboxylase and of the Coupling Factor of Photophosphorylation CF1*

The proteolytic degradation of unassembled small subunit polypeptides of ribulose-1,5-bisphosphate carboxylase and of the δ-subunit of the coupling factor of photophosphorylation CF₁ were analyzed and compared in vitro in the presence of stroma or membrane preparations from ribosome-deficient plastids isolated from 32°C-grown rye leaves (Secale cereale L.). Extracts obtained from 70S ribosome-deficient rye leaves after radioactive labeling were used as substrate source for the unassembled polypeptides. Soluble stroma as well as membrane preparations from isolated plastids contained proteolytic activities catalyzing the degradation of both the small subunits of ribulose-1,5-bisphosphate carboxylase and CF₁-δ in vitro. Maximal in vitro degradation was observed at pH 2–3 for the unassembled small subunits, but at pH 6–7 for the purified holoprotein of ribulose-1,5-bisphosphate carboxylase, and at pH 6.0 for unassembled CF₁-δ. Degradation of unassembled small subunits of ribulose-1,5-bisphosphate carboxylase at pH 3.0 was stimulated by Cu²⁺ but not by Ca²⁺, Mg²⁺ or ATP. At pH 3.0 the degradation of unassembled small subunits of ribulose-1,5-bisphosphate carboxylase was not inhibited by various protease inhibitors but was even stimulated. At pH 7.0 its degradation was inhibited by HgCl₂ and diazoacetyl nor-leucine methyl ester + Cu-acetate. The degradation of CF₁-δ was markedly inhibited by phenylmethylsulphonyl fluoride (PMSF) and to a lesser extent by 1,10-phenanthroline. According to present results different proteolytic systems appear to be involved in the degradation of unassembled small subunits of ribulose-1,5-bisphosphate carboxylase and of unassembled CF₁-δ.     

14CO2 fixation and glycolate metabolism in the dark in isolated maize (Zea mays L.) bundle sheath strands

Bundle sheath strands capable of assimilating up to 68 μmoles CO₂ per mg chlorophyll per hr in the dark have been isolated from fully expanded leaves of Zea mays L. This dark CO₂-fixing system is dependent on exogenous ribose-5-phosphate, ADP or ATP, and Mg²⁺ for maximum activity. The principal product of dark fixation in this system is 3-phosphoglycerate, indicating that the CO₂-fixing reaction is mediated by ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39). The rate of dark CO₂ uptake in the strands in the presence of saturating levels of ribose-5-phosphate plus ADP is inhibited by oxygen. The inhibitory effect of oxygen is rapidly and completely reversible, and is relieved by increased levels of CO₂. Glycolate is synthesized in this dark system in the presence of [U-¹⁴C]ribose-5-phosphate, ADP, oxygen, and an inhibitor of glycolate oxidase (EC 1.1.3.1). Glycolate formation is completely abolished by heating the strands, and the rate of glycolate synthesis is markedly reduced by either lowering the oxygen tension or increasing the level of CO₂.These results, obtained with intact cells in the absence of light, indicate that the direct inhibitory effect of oxygen on photosynthesis is associated with photosynthetic carbon metabolism, probably at the level of ribulose-1,5-bisphosphate carboxylase, and not with photophosphorylation or photosynthetic electron transport. Furthermore, the findings indicate that the synthesis of glycolate from exogenous substrate can readily occur in the absence of photosynthetic electron transport, an observation consistent with the ribulose-1, 5-bisphosphate “oxygenase” scheme for glycolate formation during photosynthesis.     

Photooxidase system of Rhodospirillum rubrum. I. Photooxidations catalyzed by chromatophores isolated from a mutant deficient in photooxidase activity

The aerobic photooxidations of reduced 2,6-dichlorophenolindophenol and of reaction-center bacteriochlorophyll (P-870) have been investigated in membrane vesicles (chromatophores) isolated from a non-phototrophic Rhodospirillum rubrum strain. In aerobic suspensions of wild-type chromatophores, continuous light elicits an increase of the levels of 2,6-dichlorophenolindophenol and of oxidized P-870, which reach steady-state values shortly after the onset of illumination. In contrast, light induces in mutant suspensions a transient increase of the levels of 2,6-dichlorophenolindophenol and of oxidized P-870, which fall to low steady-state values within a few seconds. These observations suggest that the mutation has altered a redox constituent located on the low-potential side of the photochemical reaction center, between a pool of acceptors and oxygen.Since endogenous cyclic photophosphorylation is catalyzed by mutant chromatophores at normal rates, it appears that the constituent altered by the mutation does not belong to the cyclic electron-transfer chain responsible for photophosphorylation. However, the system which mediates the aerobic photooxidations and the cyclic system are not completely independent: endogenous photophosphorylation is inhibited by oxygen in wild-type chromatophores but not in mutant chromatophores; in addition, the inhibitor of cyclic electron flow, 2-heptyl-4-hydroxyquinoline-N-oxide, enhances the aerobic photooxidation of reduced 2,6-dichlorophenolindophenol by chromatophores from both strains.These results support a tentative branched model for light-driven electron transfer. In that model, the constituent altered in the mutant strain is located in a side electron-transfer chain which connects the cyclic acceptors to oxygen.     

Ribulose-1,5-bisphosphate carboxylase-deficient plastome mutants of Oenothera

In spite of only slightly subnormal pigment contents, two plastome mutants of Oenothera (Vα, Iσ) were practically incapable of photosynthetic CO₂ fixation and another one exhibited considerably reduced photosynthesis (IVβ). While other photosynthetic enzymes were present as far as investigated, ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39) activity was very low or missing altogether. As shown by gel electrophoresis, mutant IVβ contained some, though little, fraction I protein. In the other two mutants fraction I protein could not be detected. Also, neither the small nor the large subunit of ribulose-1,5-bisphosphate carboxylase could be found in these mutants. In immunodiffusion experiments with a monospecific antiserum against rye ribulose-1,5-bisphosphate carboxylase, only extracts from wild-type Oenothera produced visible precipitation lines. Still, the presence of very low levels of immunochemically reactive antigen was indicated for all three mutants. The highest level was observed in mutant IVβ. The behaviour of the mutant extracts suggested that the antigens of mutant and wild type leaves reacting with the antiserum were not identical. All mutants appeared to have a coupled electron transport system as shown by ATP measurements, light scattering and 515 nm absorption changes. Linear electron transport was possible in the mutants. Still, the photoresponse of cytochrome f and fluorescence measurements suggested altered electron transport properties in the mutants. These are interpreted to be secondary lesions of the photosynthetic apparatus caused by primary deficiency in ribulose-1,5-bisphosphate carboxylase activity. From the absence in two mutants (Vα, Iσ) of the small subunit of ribulose-1,5-bisphosphate carboxylase, which is known to be coded for by nuclear DNA and to be synthesized on cytoplasmic ribosomes, it appears that the genetic system of the plastids is capable of interfering with the genome-controlled synthesis of plastid components.     

Hydrogen production by photosystem I of Scenedesmus: Effect of heat and salicylaldoxime on electron transport and photophosphorylation

Summary Photosynthesis, photoreduction, the p-benzoquinone Hill reaction, and glucose uptake by whole cells, as well as cyclic photophosphorylation (with PMS) by chloroplast particles were strongly inhibited by 10^-2 M salicylaldoxime or by heating whole cells for 1–2 min at 55°. In contrast, H₂ photoproduction by whole cells of mutant No. 11 and wild type Scenedesmus and PS I-mediated MR reduction by chloroplast particles were either stimulated or not significantly inhibited by these agents. H₂ production by mutant No. 8 was slightly depressed by salicylaldoxime. DCMU inhibited H₂ photoproduction with 10^-2 M salicylaldoxime approximately 20%, indicating some contribution of electrons by endogenous organic compounds to photosystem II between the O₂-evolving mechanism and the DCMU-sensitive site. We conclude that photohydrogen production by PS I of Scenedesmus does not require cyclic photophosphorylation but is due to non-cyclic electron flow from organic substrate(s) through PS I to hydrogenase where molecular H₂ is released.The following abbreviations were used CI-CCP carbonyl cyanide m-chlorophenylhydrazone - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DCPIP dichlorophenol-indophenol - MR methyl red - PMS phenazine methosulfate - PS photosystem This work was supported by contract AT-(40-1)-2687 from the U.S. Atomic Energy Commission to Professor H. Gaffron.     

Photoproduction of hydrogen by photosystem I of Scenedesmus

Summary Anaerobically adapted and illuminated Scenedesmus evolves molecular hydrogen from endogenous organic compounds. This photoproduction of H₂ does not require photosystem II, since 5x10^-6 M DCMU, which inhibited normal photosynthesis almost completely, did not significantly inhibit the photoevolution of H₂. The relative efficiencies in far-red light of photosynthesis, photoreduction and H₂ production were determined. Photohydrogen evolution was comparatively the most efficient of these three processes. Three mutants of Scenedesmus (isolated and characterized by Dr. N. I. Bishop) were also tested. Mutant PS-50, which lacks cytochrome 552, did not photoproduce H₂. Mutant No. 11, blocked in photosystem II, showed rates of H₂ production comparable to those of the wild type. Cl-CCP, an uncoupler of photophosphorylation, caused an apparent stimulation of H₂ production by mutant No. 11 and wild-type cells. Mutant No. 8, which is partially blocked in photosystem I, showed a diminished photohydrogen production which was inhibited by Cl-CCP. These results suggest that photoproduction of hydrogen by photosystem I is due either to cyclic photophosphorylation, which supplies energy needed for a dark, H₂-yielding reaction, or to a more direct photooxidation of organic compounds by the photosynthetic electron transfer chain.The following abbreviations were used: Cl-CCP=carbonyl cyanide m-chlorophenylhydrazone; DCMU=3-(3,4-dichlorophenyl)-1,1-dimethylurea.This work was supported by contract AT-(40-1)-2687 from the U.S. Atomic Energy Commission to Professor H. Gaffron.     

Antimycin A Stimulation of Rate-limiting Steps of Photosynthesis in Isolated Spinach Chloroplasts

Changes in levels of metabolites in isolated spinach (Spinacia oleracea) chloroplasts seen upon addition of antimycin A suggest that the activities of enzymes mediating several regulated reactions are affected. Apparently, the presence of added antimycin A does not increase the level of CO₂ in the chloroplasts, nor does it stimulate CO₂ fixation by increasing the level of the carboxylation substrate, ribulose-1,5-diphosphate. Rather, it appears that antimycin A increases CO₂ fixation rate by indirectly stimulating the enzyme, ribulose-1,5-diphosphate carboxylase (E.C. 4.1.1.39), which mediates the carboxylation of ribulose-1,5-diphosphate to give 3-phosphoglycerate. Another rate-limiting enzyme of the reductive pentose phosphate cycle, hexose diphosphatase (E.C. 3.1.3.11), seems also to be stimulated. The synthesis of polysaccharides (mostly starch) seems also to be stimulated. These results are interpreted as indicating that antimycin A addition enhances the general activation of those enzymes which already are activated during photosynthesis but are inactive in the dark. The ratio of adenosine triphosphate-adenosine diphosphate under conditions of photosynthesis was only moderately decreased in the presence of antimycin A, perhaps accounting in part for an observed increase in accumulation of 3-phosphoglycerate as compared with dihydroxyacetone phosphate. No significant effect on movement of metabolites from the chloroplast to the medium was seen.     

CHLOROPLAST STRUCTURE AND FUNCTION IN ac-20, A MUTANT STRAIN OF CHLAMYDOMONAS REINHARDI : I. CO2 Fixation and Ribulose-1,5-Diphosphate Carboxylase Synthesis

A mutant strain of the green alga Chlamydomonas reinhardi, ac-20, is described in which both the rate of CO₂ fixation by whole cells and the rate of carboxylation of ribulose-1,5-diphosphate in cell-free extracts are reduced, particularly when sodium acetate is present in the growth medium. Of the enzymes of the reductive pentose phosphate cycle tested, only ribulose-1,5-diphosphate carboxylase activity is reduced in the mutant strain, and it appears that the low carboxylase activity limits the strain's rate of photosynthetic carbon metabolism. Evidence is presented to show that the fluctuation in the level of the enzyme activity in the presence or absence of acetate results from the fluctuation in the level of some factor(s) limiting the rate of synthesis of the protein.     

Photosynthetic Activity of Chloroplasts Isolated From Bermudagrass (Cynodon dactylon L.), a Species With a High Photosynthetic Capacity

Chloroplasts have been isolated from bermudagrass (Cynodon dactylon L.) leaves and assayed for photophosphorylation and electron transport activity. These chloroplasts actively synthesize adenosine triphosphate during cyclic electron flow with phenazine methosulfate and noncyclic electron flow concurrent with the reduction of such Hill oxidants as nicotinamide adenosine dinucleotide phosphate, cytochrome c, and ferricyanide. Apparent Km values for the cofactors of photophosphorylation have been determined to be 5 × 10⁻⁵ M for phosphate and 2.5 × 10⁻⁵ M for adenosine diphosphate. The influence of light intensity on photophosphorylation has been studied and the molar ratio of cyclic to noncyclic phosphorylation calculated. It is concluded that the high photosynthetic capacity of bermudagrass leaves probably could be supported by the photophosphorylation capacities indicated in these chloroplast studies and the anomalous lack of data in chlorolast studies on the production of sufficient reductant for CO₂ assimilation at high light intensities has been noted.     

In vitro and in vivo analyses of the role of the carboxysomal β-type carbonic anhydrase of the cyanobacterium Synechococcus elongatus in carboxylation of ribulose-1,5-bisphosphate

The carboxylase activities of crude carboxysome preparations obtained from the wild-type Synechococcus elongatus strain PCC 7942 strain and the mutant defective in the carboxysomal carbonic anhydrase (CA) were compared. The carboxylation reaction required high concentrations of bicarbonate and was not even saturated at 50 mM bicarbonate. With the initial concentrations of 50 mM and 25 mM for bicarbonate and ribulose-1,5-bisphosphate (RuBP), respectively, the initial rate of RuBP carboxylation by the mutant carboxysome (0.22 μmol mg^?1 protein min^?1) was only 30 % of that observed for the wild-type carboxysomes (0.71 μmol mg^?1 protein min^?1), indicating the importance of the presence of CA in efficient catalysis by ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco). While the mutant defective in the ccmLMNO genes, which lacks the carboxysome structure, could grow under aeration with 2 % (v/v) CO₂ in air, the mutant defective in ccaA as well as ccmLMNO required 5 % (v/v) CO₂ for growth, indicating that the cytoplasmically localized CcaA helped utilization of CO₂ by the cytoplasmically localized Rubisco by counteracting the action of the CO₂ hydration mechanism. The results predict that overexpression of Rubisco would hardly enhance CO₂ fixation by the cyanobacterium at CO₂ levels lower than 5 %, unless Rubisco is properly organized into carboxysomes.     

Interactions between ribulose-1,5-bisphosphate carboxylase and stromal metabolites. II. Corroboration of the role of this enzyme as a metabolite buffer

The capacity of ribulose-1,5-bisphosphate carboxylase to bind reversibly chloroplast metabolites which are the substrates for both thylakoid and stromal enzymes was assessed using spinach chloroplasts and chloroplast extracts and with pure wheat ribulose-1,5-bisphosphate carboxylase. Measurements of the rate of coupled electron flow to methyl viologen in ‘leaky’ chloroplasts (which retained the chloroplast envelope and stromal enzymes but which were permeable to metabolites) and also with broken chloroplasts and washed thylakoids were used to study the effects of binding ADP and inorganic phopshate to ribulose-1,5-bisphosphate carboxylase. The presence of ribulose-1,5-bisphosphate carboxylase significantly altered the values obtained for apparent K_m for inorganic phosphate and ADP of coupled electron transport. The K_m (P_i) in washed thylakoids was 60–80 μM, in ‘leaky’ chloroplasts it was increased to 180–200 μM, while in ‘leaky’ chloroplasts preincubated with KCN and ribulose 1,5-bisphosphate the value was decreased to 40–50 μM. Similarly, the K_m (ADP) of coupled electron transport in washed thylakoids was 60–70 μM, in ‘leaky’ chloroplasts it was 130–150 μM and with ‘leaky’ chloroplasts incubated in the presence of KCN and ribulose 1,5-bisphosphate a value of 45–50 μM was obtained. The ability of ribulose 1,5-bisphosphate carboxylase to reduce the levels of free glycerate 3-phosphate in the absence of ribulose 1,5-bisphosphate was examined using a chloroplast extract system by varying the concentrations of stromal protein or purified ribulose 1,5-bisphosphate carboxylase. The effect of binding glycerate 3-phosphate to ribulose-1,5-bisphosphate carboxylase on glycerate 3-phosphate reduction was to reduce both the rate an the amount of NADPH oxidation for a given amount of glycerate 3-phosphate added. The addition of ribulose 1,5-bisphosphate reinitiated NADPH oxidation but ATP or NADPH did not. Incubation of purified ribulose-1,5-bisphosphate carboxylase with carboxyarabinitolbisphosphate completely inhibited the catalytic activity of the enzyme and decreased inhibition of glycerate-3-phosphate reduction. Two binding sites with different affinities for glycerate 3-phosphate were observed with pure ribulose-1,5-bisphosphate carboxylase.     

Effects of calcium on the photosynthesis of intact leaves and isolated chloroplasts of sugar beets

Effects of calcium on photosynthesis in sugar beets (Beta vulgaris L. cv. F58-554H1) were studied by inducing calcium deficiency and determining changes in CO₂ uptake by attached leaves, electron transport, and photophosphorylation by isolated chloroplasts, and CO₂ assimilation by ribulose diphosphate carboxylase extracts. Calcium deficiency had no significant effect on leaf CO₂ uptake, photoreduction of ferricyanide, cyclic or noncyclic ATP formation of isolated chloroplasts, or on ribulose diphosphate carboxylase CO₂ assimilation, when the rates were expressed per unit chlorophyll. When expressed per unit leaf area CO₂ uptake increased by about 15% in low calcium leaves. The most noticeable effect of calcium deficiency was reduction in leaf area: low calcium had no effect on dark respiratory CO₂ evolution, on leaf diffusion resistance, or on mesophyll resistance to CO₂. We concluded that only small amounts of calcium are required for normal photosynthetic activity of sugar beet leaves.     

An improved model of C3 photosynthesis at high CO2: Reversed O2 sensitivity explained by lack of glycerate reentry into the chloroplast

Current models of C₃ photosynthesis incorporate a phosphate limitation to carboxylation which arises when the capacity for starch and sucrose synthesis fails to match the capacity for the production of triose phosphates in the Calvin cycle. As a result, the release of inorganic phosphate in the chloroplast stroma fails to keep pace with its rate of sequestration into triose phosphate, and phosphate becomes limiting to photosynthesis. Such a model predicts that when phosphate is limiting, assimilation becomes insensitive to both CO₂ and O₂, and is thus incapable of explaining the experimental observation that assimilation, under phosphate-limited conditions, frequently exhibits reversed sensitivity to both CO₂ and O₂, i.e., increasing O₂ stimulates assimilation and increasing CO₂ inhibits assimilation. We propose a model which explains reversed sensitivity to CO₂ and O₂ by invoking the net release of phosphate in the photorespiratory oxidation cycle. In order for this to occur, some fraction of the glycollate carbon which leaves the stroma and which is recycled to the chloroplast by the photorespiratory pathway as glycerate must remain in the cytosol, perhaps in the form of amino acids. In that case, phosphate normally used in the stromal glycerate kinase reaction to generate PGA from glycerate is made available for photophosphorylation, stimulating RuBP regeneration and assimilation. The model is parameterized for data obtained on soybean and cotton, and model behavior in response to CO₂, O₂, and light is demonstrated.Abbreviations PFD photon flux density - PGA 3-phosphoglycerate - Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase - RuBP ribulose-1,5-bisphosphate - TPU triose phosphate utilization     

Operation of the glycolate pathway in isolated bundle sheath strands of maize and Panicum maximum

Operation of the glycolate pathway in isolated bundle sheath (BS) strands of two C₄ species was demonstrated from ¹⁴C incorporation into two intermediates, glycine and serine, under conditions favourable for photorespiratory activity. Isolated BS strands fixing ¹⁴CO₂ under light at physiological rates incorporate respectively 3% (Zea mays L., cv. INRA 258) and 7% (Panicum maximum Jacq.) of total ¹⁴C fixed into glycine + serine, at low bicarbonate levels (less than the Km for CO₂ fixation, 0.8 mM). Higher bicarbonate concentrations depressed the percentage of incorporation into the two amino acids. No labelling was observed in the absence of added glutamate. Oxygen was required for glycine + serine labelling, since ¹⁴C incorporation into glycine was largely depressed by argon flushing, and labelling of the two amino acids was nearly suppressed by the addition of the strong reductant, dithionite, especially in maize. Two inhibitors of the glycolate pathway were tested. With α-hydroxypyridine-methanesulfonic acid, an inhibitor of glycolate oxidase, labelling of glycine and serine remained minimal whereas glycolate was accumulated. Isoniazid, an inhibitor of the transformation of glycine to serine induced a 50% increased labelling of glycine in maize BS, and a large decrease in serine labelling. In Panicum, the increase in [¹⁴C]-glycine was 90%. These results suggest that the pathway glycolate → glycine → serine operates in these plants. However, leakage of metabolites occurs in BS cells, especially in maize and a large part of newly formed glycolate, glycine and serine is exported out of the cells. Operation of ribulose-1,5-bisphosphate oxygenase activity in competition with ribulose-1,5-bisphosphate carboxylase is demonstrated by the lowering of total ¹⁴CO₂ fixation when O₂ is increased at low bicarbonate concentration. An interesting feature observed in maize BS, at low bicarbonate concentration, was an increase in ribulose-1,5-bisphosphate labelling when the O₂ level was decreased. This was accompanied by an increase in CO₂ fixation. This could indicate an increased rate in synthesis of ribulose-1,5-bisphosphate (which accumulated) due to a stimulation of ATP synthesis by cyclic photophosphorylation under anaerobic conditions.     

Leaf catalase mRNA and catalase-protein levels in a high-catalase tobacco mutant with o(2)-resistant photosynthesis

Experiments were conducted with a tobacco (Nicotiana tabacum) mutant with 40 to 50% greater catalase activity than wild type that is associated with a novel form of O₂-resistant photosynthesis. The apparent K_m for H₂O₂ was the same in mutant and wild-type leaf extracts. Tobacco RNAs were hybridized with Nicotiana sylvestris catalase cDNA, and a threefold greater steady-state level of catalase mRNA was found in mutant leaves. Steady-state levels of ribulose-1,5-bisphosphate carboxylase small subunit mRNA were similar in mutant and wild type. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of partially purified catalase showed that the protein concentration in the band corresponding to catalase was higher in the mutant than in the wild type. Separation of leaf catalase proteins by isoelectric focusing revealed the presence of five major bands and one minor band of activity. The distribution of the catalase activity among these forms was similar in mutant and wild type, although the total activity was higher in the mutant in all five major bands. The results indicate that the enhanced catalase activity in mutant leaves is caused by an increase in synthesis of catalase protein and that this trait is mediated at the nucleic acid level.     

Cyclic photophosphorylation by chromatophores of the facultative phototroph,Rhodopseudomonas capsulata

Summary Cyclic photophosphorylation catalyzed by chromatophores derived from the facultative phototroph, Rhodopseudomonas capsulata was investigated. In the absence of an external electron donor such as succinate, cyclic photophosphorylation is strongly inhibited by O₂. Maximal phosphorylation rates are obtained in the presence of molecular hydrogen. Cytochrome c and bovine serum albumin have no significant effects on the reaction. However, dichlorophenolindophenol and phenazonium methosulfate are inhibitory to cyclic photophosphorylation. Cyclic photophosphorylation is sensitive to antimycin A, but highly resistant to heptylhydroxy-quinoline-N-oxide. Neither phenazonium methosulfate, nor dichlorophenolindophenol or tetramethyl-p-phenylenediamine can effect antimycin-insensitive cyclic photophosphorylation. Oligomycin strongly inhibits the phosphorylation. Overreduction caused by the ascorbate-dichlorophenolindophenol couple results in strong inhibition of phosphorylation. Addition of fumarate decreases the inhibition caused by overreduction. However, the fumarate mediated phosphorylation is nearly completely inhibited by antimycin A. Atebrine is a strong inhibitor for cyclic photophosphorylation, whereas dinitrophenol is only a weak inhibitor.

---

Zusammenfassung Die durch Chromatophoren aus dem fakultativ phototrophen Rhodopseudomonas capsulata katalysierte cyclische Photophosphorylierung wurde untersucht. In der Abwesenheit eines zusätzlichen Elektronendonators wie Succinat wird die cyclische Photophosphorylierung durch O₂ stark gehemmt. Maximale Phosphorylierungsraten werden unter H₂-Atmosphäre erzielt. Cytochrom c und Rinderserumalbumin haben keinen deutlichen Effekt auf die Reaktion. Demgegenüber haben Dichlorphenolindophenol und Phenazinmethosulfat eine hemmende Wirkung auf die cyclische Photophosphorylierung. Die cyclische Photophosphorylierung wird durch Antimycin A stark gehemmt, ist aber gegenüber Heptyl-hydroxy-chinolin-N-oxyd auffallend resistent. Weder Phenazinmethosulfat noch Dichlorphenolindophenol oder Tetramethyl-p-phenylendiamin bewirken eine antimycin-resistente Phosphorylierung. Oligomycin hemmt die Photophosphorylierung stark. Eine durch Ascorbat-Dichlorphenolindophenol verursachte Überreduktion wirkt sich stark hemmend auf die Phosphorylierung aus. In Gegenwart von Fumarat ist die durch Überreduktion bedingte Hemmung stark verringert. Diese vom Fumarat abhängige Photophosphorylierung wird jedoch durch Antimycin A beinahe vollständig gehemmt. Atebrin ist ein starker Hemmstoff für die cyclische Photophosphorylierung. Demgegenüber ist die durch Dinitrophenol bewirkte Hemmung der cyclischen Photophosphorylierung gering.

---

Abbreviations ADP adenosine diphosphate - ATP adenosine triphosphate - BChl bacteriochlorophyll - DNP 2,4-dinitrophenol - DCPIP dichlorophenolindophenol - FAD flavinadenine dinucleotide - FMN flavin mononucleotide - G-6-P glucose-6-phosphate - HOQNO heptylhydroxy-quinoline-N-oxide - NAD(P) nicotinamid-adenine-dinucleotide (phosphate) - PMS phenazonium methosulfate - Rh. Rhodospirillum - Rhps. Rhodopseudomonas - TMPD tetramethyl-p-phenylenediamine