Ribulose 1,5-diphosphate carboxylase and Chlorobium thiosulfatophilum

1. Cell-free extracts of the photosynthetic bacterium Chlorobium thiosulfatophilum, strains 8327 and Tassajara, were assayed for ribulose 1,5-diphosphate (RuDP) carboxylase and phosphoribulokinase-the two enzymes peculiar to the reductive pentose phosphate cycle. 2. RuDP carboxylase was consistently absent in strain 8327. The Tassajara strain showed a low RuDP-dependent CO₂ fixation activity that was somewhat higher in cells following transatlantic air shipment than in freshly grown cells. The stability and behaviour of this activity in sucrose density gradients were similar to those described by other workers. 3. The radioactive carboxylation products formed in the presence of RuDP by enzyme preparations from the Tassajara strain did not include 3-phosphoglycerate-the known product of the RuDP carboxylase reaction, but instead consisted of the unrelated acids glutamate, aspartate and malate. 4. Phosphoribulokinase was absent in all preparations of the two Chlorobium strains tested. By contrast, phosphoribulokinase as well as RuDP carboxylase were readily demonstrated in preparations from pea chloroplasts and the photosynthetic bacterium Rhodospirillum rubrum. 5. It is concluded that C. thiosulfatophilum appears to lack RuDP carboxylase, phosphoribulokinase, and hence, the reductive pentose phosphate cycle.Support of a J. S. Guggenheim Fellowship is gratefully acknowledged     

Photosynthetic/photorespiratory characteristics of C3−C4 intermediate species

The extent of photorespiration, the inhibition of apparent photosynthesis (APS) by 21% O₂, and the leaf anatomical and ultrastructural features of the naturally occurring C₃–C₄ intermediate species in the diverse Panicum, Moricandia, and Flaveria genera are between those features of representative C₃ and C₄ plants. The greatest differences between the photosynthetic/photorespiratory CO₂ exchange characteristics of the C₃–C₄ intermediates and C₃ plants occur for the parameters which are measured at low pCO₂ (i.e., the CO₂ compensation concentration and rates of CO₂ evolution into CO₂-free air in the light). The rates of APS by the intermediate species at atmospheric pCO₂ are similar to those of C₃ plants.The mechanisms which are responsible for reducing photorespiration in the C₃–C₄ intermediate species are poorly understood, but two proposals have been advanced. One emphasizes the importance of limited C₄ photosynthesis which reduces O₂ fixation by ribulose 1,5-bisphosphate carboxylase/oxygenase, and, thus, reduces photorespiration by a CO₂-concentrating mechanism, while the other emphasizes the importance of the internal recycling of photorespiratory CO₂ evolved from the chloroplast/mitochondrion-containing bundle-sheath cells. There is no evidence from recent studies that limited C₄ photosynthesis is responsible for reducing photorespiration in the intermediate Panicum and Moricandia species. However, preliminary results suggest that some, but not all, of the intermediate Flaveria species may possess a limited C₄ cycle. The importance of a chlorophyllous bundle-sheath layer in the leaves of intermediate Panicum and Moricandia species in a mechanism based on the recycling of photorespiratory CO₂ is uncertain.Therefore, although they have yet to be clearly delineated, different strategies appear to exist in the C₃–C₄ intermediate group to reduce photorespiration. Of major importance is the finding that some mechanism(s) other than Crassulacean acid metabolism or C₄ photosynthesis has (have) evolved in at least the majority of these terrestrial intermediate species to reduce the seemingly wasteful metabolic process of photorespiration.Abbreviations APS apparent (net) photosynthesis - CAM Crassulacean acid metabolism - CE carboxylation efficiency - T CO₂ compensation concentration - IRGA infrared gas analysis - P_i orthophosphate - PEP phosphoenolpyruvate - RuBP ribulose 1,5-bisphosphate Published as Paper No. 7383, Journal Series, Nebraska Agricultural Experiment Station.     

Regulation of nitrogenase activity by covalent modification in Chromatium vinosum

Nitrogenase in Chromatium vinosum was rapidly, but reversibly inhibited by NH ₄ ⁺ . Activity of the Fe protin component of nitrogenase required both Mn²⁺ and activating enzyme. Activating enzyme from Rhodospirillum rubrum could replace Chromatium chromatophores in activating the Chromatium Fe protein, and conversely, a protein fraction prepared from Chromatium chromatophores was effective in activating R. rubrum Fe protein. Inactive Chromatium Fe protein contained a peptide covalently modified by a phosphate-containing molecule, which migrated the same in SDS-polyacrylamide gels as the modified subunit of R. rubrum Fe protein. In sum, these observations suggest that Chromatium nitrogenase activity is regulated by a covalent modification of the Fe protein in a manner similar to that of R. rubrum.Abbreviation HEPES N-2-hydroxyethyl piperazine-N-2-ethanesulfonic acid     

Effect of dissolved inorganic carbon on the expression of carboxysomes,localization of Rubisco and the mode of inorganic carbon transport in cells of the cyanobacterium Synechococcus UTEX 625

In the cyanobacterium Synechococcus UTEX 625, the extent of expression of carboxysomes appeared dependent on the level of inorganic carbon (CO₂+HCO _inf3 ^sup- ) in the growth medium. In cells grown under 5% CO₂ and in those bubbled with air, carboxysomes were present in low numbers (<2 · longitudinal section^-1) and were distributed in an apparently random manner throughout the centroplasm. In contrast, cells grown in standing culture and those bubbled with 30 l CO₂ · 1^-1 possessed many carboxysomes (>8 · longitudinal section^-1). Moreover, carboxysomes in these cells were usually positioned near the cell periphery, aligned along the interface between the centroplasm and the photosynthetic thylakoids. This arrangement of carboxysomes coincided with the full induction of the HCO _inf3 ^sup- transport system that is involved in concentrating inorganic carbon within the cells for subsequent use in photosynthesis. Immunolocalization studies indicate that the Calvin cycle enzyme ribulose bisphosphate carboxylase was predominantly carboxysome-localized, regardless of the inorganic carbon concentration of the growth medium, while phosphoribulokinase was confined to the thylakoid region. It is postulated that the peripheral arrangement of carboxysomes may provide for more efficient photosynthetic utilization of the internal inorganic carbon pool in cells from cultures where carbon resources are limiting.Abbreviations Chl chlorophyll - DIC dissolved inorganic carbon (CO₂+HCO _inf3 ^sup- +CO _inf3 ^sup2- ) - PRK phosphoribulokinase - RuBP ribulose 1,5-bisphosphate - Rubisco LS large subunit of ribulose 1,5-bisphosphate carboxylase/oxygenase     

Modelling C3 photosynthesis: A sensitivity analysis of the photosynthetic carbon-reduction cycle

A model of the C ₃ photosynthetic system is developed which describes the sensitivity of the steadystate rate of carbon dioxide assimilation to changes in the activity of several enzymes of the system. The model requires measurements of the steady-state rate of carbon dioxide assimilation, the concentrations of several intermediates in the photosynthetic system, and the concentration of the active site of ribulose 1,5-bisphosphate carboxyalse/oxygenase (Rubisco). It is shown that in sunflowers (Helianthus annuus L.) at photon flux densities that are largely saturating for the rate of photosynthesis, the steady-stete rate of carbon dioxide assimilation is most sensitive to Rubisco activity and, to a lesser degree, to the activities of the stromal fructose, 6-bisphosphatase and the enzymes catalysing sucrose synthesis. The activities of sedoheptulose 1,7-bisphosphatase, ribulose 5-phosphate kinase, ATP synthase and the ADP-glucose pyrophosphorylase are calculated to have a negligible effect on the flux under the high-light conditions. The utility of this analysis in developing simpler models of photosynthesis is also discussed.Abbreviations c _i intercellular CO₂ concentration - C _infP ^supJ control coefficient for enzyme P with respect to flux J - DHAP dihydroxyacetonephosphate - E4P erythrose 4-phosphate - F6P fructose 6-phosphate - FBP fructose 1,6-bisphosphate - FBPase fructose 1,6-bisphosphatase - G3P glyceraldehyde 3-phosphate - G1P glucose 1-phosphate - G6P glucose 6-phosphate - Pi inorganic phosphate - PCR photosynthetic carbon reduction - PGA 3-phosphoglyceric acid - PPFD photosynthetically active photon flux density - R _n ^J response coefficient for effector n with respect to flux J - R5P ribose 5-phosphate - Rubisco ribulose 1,5-bisphosphate carboxylase/oxygenase - Ru5P ribulose 5-phosphate - RuBP ribulose 1,5-bisphosphate - S7P sedoheptulose 7-phosphate - SBP sedoheptulose 1,7-bisphosphate - SBPase sedoheptulose 1,7-bisphosphatase - SPS sucrose-phosphate synthase - Xu5P xylulose 5-phosphate - _n ^P elasticity coefficient for effector n with respect to the catalytic velocity of enzyme P This research was funded by an Australian Research Council grant to I.E.W. and was undertaken during a visity by K.A.M. to the James Cook University of North Queensland. The expert help of Glenys Hanley and Mick Kelly is greatly appreciated.     

Autotrophic growth of four Sulfolobus strains on tetrathionate and the effect of organic nutrients

Autotrophic growth yields of four strains of Sulfolobus using tetrathionate as sole energy substrate fell in the range 6.2–7.8 g dry weight (mol tetrathionate oxidized)^-1. Autotrophic organisms lacked ribulose 1,5-bis-phosphate carboxylase, but contained pyruvate and phosphoenolpyruvate carboxylases. S. brierleyi and strains B6-2 and LM exhibited mixotrophic growth, with tetrathionate oxidation, CO₂-fixation and organic substrate assimilation occurring concurrently, using media containing glucose or acetate. Yeast extract or succinate supported heterotrophic growth and showed strain-dependent repression of one or both of tetrathionate oxidation and CO₂-fixation resulting in biphasic growth. All four carbon atoms of succinate were assimilated to cell-carbon during growth. Acetate was the major source of cell-carbon during mixotrophic growth. These observations are not inconsistent with the possibility of a reductive carboxylic acid cycle in these organisms. Radiorespirometric analysis of glucose oxidation indicated CO₂ release to occur by means of an Entner-Doudoroff pathway (followed by pyruvate decarboxylation) and oxidative pentose phosphate pathway reactions. There was little evidence from the glucose radiorespirometry of the large-scale use of an oxidative tricarboxylic acid cycle for terminal oxidation of acetate derived from pyruvate. These results demonstrate the considerable metabolic versatility of Sulfolobus strains and show that there is significant variation among them.Abbreviations PIPES Piperazine-N,N-bis (2-ethane sulphonic acid)     

Optimal acclimation of the C3 photosynthetic system under enhanced CO2

A range of studies of C₃ plants have shown that there is a change in both the carbon flux and the pattern of nitrogen allocation when plants are grown under enhanced CO₂. This paper examines evidence that allocation of nitrogen both to and within the photosynthetic system is optimised with respect to the carbon flux. A model is developed which predicts the optimal relative allocation of nitrogen to key enzymes of the photosynthetic system as a function of CO₂ concentration. It is shown that evidence from flux control analysis is broadly consistent with this model, although at high nitrogen and under certain conditions at low nitrogen experimental data are not consistent with the model. Acclimation to enhanced CO₂ is also assessed in terms of resource allocation between photosynthate sources and sinks. A means of assessing the optimisation of this source-sink allocation is proposed, and several studies are examined within this framework. It is concluded that C₃ plants probably possess the genetic feedback mechanisms required to efficiently smooth out any imbalance within the photosynthetic system caused by a rise in atmospheric CO₂.Abbreviations A net rate of CO₂ assimilation - c _i intercellular CO₂ concentration - C_R ^A flux control coefficient for Rubisco with respect to flux A - FBPase fructose 1,6-bisphosphatase - k_app apparent catalytic rate constant - PCO photorespiratory carbon oxidation - PCR photosynthetic carbon reduction - PPFD photosynthetically active photon flux density - Rubisco ribulose 1,5-bisphosphate carboxylase/oxygenase - RuBP ribulose 1,5-bisphosphate - Ru5P ribulose 5-phosphate - SBPase sedoheptulose 1,7-bisphosphatase     

Autotrophic growth and CO2 fixation of Chloroflexus aurantiacus

Chlorofluexus aurantiacus OK-70 fl was grown photoautotrophically with hydrogen as the electron source. The lowest doubling time observed was 26 h.The mechanism of CO₂ fixation in autotrophically grown cells was studied. The presence of ribulose-1,5-bis-phosphate carboxylase and phosphoribulokinase could not be demonstrated. Carbon isotope fractionation (¹³C) was small, and alanine and aspartate but not 3-phosphoglycerate were the major labelled compounds in short term ¹⁴CO₂ labelling. Thus CO₂ is not fixed by the Calvin cycle.Fluoroacetate (FAc) completely inhibited protein synthesis in cultures and caused a slight citrate accumulation. However, CO₂ fixation continued and increased polyglucose formation occurred. Under these conditions added acetate was metabolized to polyglucose, as were glycine, serine, glyoxylate and succinate, but to a lesser extent; little or no formate or CO was utilised.Glyoxylate inhibited CO₂ fixation in vivo, indicating that pyruvate is formed from acetyl-CoA and CO₂ by pyruvate synthase. Two key enzymes of the reductive TCA cycle, citrate lyase and -ketoglutarate synthase were not detected in cell free extracts, but pyruvate synthase and phosphoenolpyruvate carboxylase were demonstrated. It is concluded that acetyl-CoA is a central intermediate in the CO₂ fixation process, but the mechanism of its synthesis is not clear.Abbreviations Rubisco ribulose-1,5-bisphosphate carboxylase - TCA cycle tricarboxylic acid cycle - FAc monofluoroacetate - PEP phosphoenolpyruvate - MV methyl viologen - TTC triphenyltetrazolium chloride - PMS phenazine methosulfate     

Photorespiratory metabolism and immunogold localization of photorespiratory enzymes in leaves of C3 and C3-C4 intermediate species of Moricandia

Photorespiratory metabolism of the C₃-C₄ intermediate species Moricandia arvensis (L.) DC has been compared with that of the C₃ species, Moricandia moricandioides (Boiss.) Heywood. Assays of glycollate oxidase (EC 1.1.3.1), glyoxylate aminotransferases (EC 2.6.1.4, EC 2.6.1.45) and hydroxypyruvate reductase (EC 1.1.1.29) indicate that the capacity for flux through the photorespiratory cycle is similar in both species. Immunogold labelling with monospecific antibodies was used to investigate the cellular locations of ribulose 1,5-bisphosphate carboxylase/oxygenase (EC 4.1.1.39), glycollate oxidase, and glycine decarboxylase (EC 2.1.2.10) in leaves of the two species. Ribulose 1,5-bisphosphate carboxylase/oxygenase was confined to the stroma of chloroplasts and glycollate oxidase to the peroxisomes of all photosynthetic cells in leaves of both species. However, whereas glycine decarboxylase was present in the mitochondria of all photosynthetic cells in M. moricandioides, it was only found in the mitochondria of bundle-sheath cells in M. arvensis. We suggest that localized decarboxylation of glycine in the leaves of M. arvensis will lead to improved recapture of photorespired CO₂ and hence a lower rate of photorespiration.Abbreviations kDa kilodalton - RuBP ribulose-1,5-bisphosphate     

Co-function of C3-and C4-photosynthetic pathways in C3, C4 and C3-C4 intermediate Flaveria species

The potential for C₄ photosynthesis was investigated in five C₃-C₄ intermediate species, one C₃ species, and one C₄ species in the genus Flaveria, using ¹⁴CO₂ pulse-¹²CO₂ chase techniques and quantum-yield measurements. All five intermediate species were capable of incorporating ¹⁴CO₂ into the C₄ acids malate and aspartate, following an 8-s pulse. The proportion of ¹⁴C label in these C₄ products ranged from 50–55% to 20–26% in the C₃-C₄ intermediates F. floridana Johnston and F. linearis Lag. respectively. All of the intermediate species incorporated as much, or more, ¹⁴CO₂ into aspartate as into malate. Generally, about 5–15% of the initial label in these species appeared as other organic acids. There was variation in the capacity for C₄ photosynthesis among the intermediate species based on the apparent rate of conversion of ¹⁴C label from the C₄ cycle to the C₃ cycle. In intermediate species such as F. pubescens Rydb., F. ramosissima Klatt., and F. floridana we observed a substantial decrease in label of C₄-cycle products and an increase in percentage label in C₃-cycle products during chase periods with ¹²CO₂, although the rate of change was slower than in the C₄ species, F. palmeri. In these C₃-C₄ intermediates both sucrose and fumarate were predominant products after a 20-min chase period. In the C₃-C₄ intermediates, F. anomala Robinson and f. linearis we observed no significant decrease in the label of C₄-cycle products during a 3-min chase period and a slow turnover during a 20-min chase, indicating a lower level of functional integration between the C₄ and C₃ cycles in these species, relative to the other intermediates. Although F. cronquistii Powell was previously identified as a C₃ species, 7–18% of the initial label was in malate+aspartate. However, only 40–50% of this label was in the C-4 position, indicating C₄-acid formation as secondary products of photosynthesis in F. cronquistii. In 21% O₂, the absorbed quantum yields for CO₂ uptake (in mol CO₂·[mol quanta]^-1) averaged 0.053 in F. cronquistii (C₃), 0.051 in F. trinervia (Spreng.) Mohr (C₄), 0.052 in F. ramosissima (C₃-C₄), 0.051 in F. anomala (C₃-C₄), 0.050 in F. linearis (C₃-C₄), 0.046 in F. floridana (C₃-C₄), and 0.044 in F. pubescens (C₃-C₄). In 2% O₂ an enhancement of the quantum yield was observed in all of the C₃-C₄ intermediate species, ranging from 21% in F. ramosissima to 43% in F. pubescens. In all intermediates the quantum yields in 2% O₂ were intermediate in value to the C₃ and C₄ species, indicating a co-function of the C₃ and C₄ cycles in CO₂ assimilation. The low quantum-yield values for F. pubescens and F. floridana in 21% O₂ presumably reflect an ineffcient transfer of carbon from the C₄ to the C₃ cycle. The response of the quantum yield to four increasing O₂ concentrations (2–35%) showed lower levels of O₂ inhibition in the C₃-C₄ intermediate F. ramosissima, relative to the C₃ species. This indicates that the co-function of the C₃ and C₄ cycles in this intermediate species leads to an increased CO₂ concentration at the site of ribulose-1,5-bisphosphate carboxylase/oxygenase and a concomitant decrease in the competitive inhibition by O₂.Abbreviations PEP phosphoenolpyruvate - PGA 3-phosphoglycerate - RuBP ribulose-1,5-bisphosphate     

Growth,14CO2 fixation,activites of photosystems,ribulose 1,5-bisphosphate carboxylase and nitrate reductase in trees as affected by simulated acid rain

In seedlings of the tropical tree speciesErythrina variegata Lam. andHardwickia binata Roxb. exposed to different acidic mist (H₂SO₄, pH 5, 3 and 2) for 5 d significant reduction in seedling growth, biomass accumulation and¹⁴CO₂ fixation were determined. In isolated chloroplasts a decrease in the activities of photosystem 2 and whole electron transport chain was observed only at pH 3 and 2, but no significant change in photosystem 1 activity was observed. SDS-PAGE analysis of crude leaf extracts of ribulose 1,5-bisphosphate carboxylase (RuBPC) indicated a significant loss of 55 and 15 kDa polypeptides at pH 2 inErythrina. The reduction in the RuBPC activity in seedlings grown under acidic mists correlated well with CO₂ fixation.     

How various factors influence the CO2/O2 specificity of ribulose-1,5-bisphosphate carboxylase/oxygenase

Temperature, activating metal ions, and amino-acid substitutions are known to influence the CO₂/O₂ specificity of the chloroplast enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase. However, an understanding of the physical basis for enzyme specificity has been elusive. We have shown that the temperature dependence of CO₂/O₂ specificity can be attributed to a difference between the free energies of activation for the carboxylation and oxygenation partial reactions. The reaction between the 2,3-enediolate of ribulose 1,5-bisphosphate and O₂ has a higher free energy of activation than the corresponding reaction of this substrate with CO₂. Thus, oxygenation is more responsive to temperature than carboxylation. We have proposed possible transition-state structures for the carboxylation and oxygenation partial reactions based upon the chemical natures of these two reactions within the active site. Electrostatic forces that stabilize the transition state of the carboxylation reaction will also inevitably stabilize the transition state of the oxygenation reaction, indicating that oxygenase activity may be unavoidable. Furthermore, the reduction in CO₂/O₂ specificity that is observed when activator Mg²⁺ is replaced by Mn²⁺ may be due to Mg²⁺ being more effective in neutralizing the negative charge of the carboxylation transition state, whereas Mn²⁺ is a transition-metal ion that can overcome the triplet character of O₂ to promote the oxygenation reaction.Abbreviations CABP 2-carboxyarabinitol 1,5-bisphosphate - enol-RuBP 2,3-enediolate of ribulose 1,5-bisphosphate - K_c K_mfor CO₂ - K_o K_mfor O₂ - Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase - RuBP ribulose 1,5-bisphosphate - V_c V _max for carboxylation - V_o V _max for oxygenation     

High CO2 partial pressure effects on dark and light CO2 fixation and metabolism in Vicia faba leaves

Stomatal opening on Vicia faba can be induced by high CO₂ partial pressures (10.2%) in dark as well as in light. Stomatal aperture was measured in both cases with a hydrogen porometer. The distribution of ¹⁴C among early products of photosynthesis was studied. Comparisons are made with carboxylations occurring when stomata were open in the dark with CO₂-free air and in light with 0.034% CO₂. Results showed that in high CO₂ partial pressure in light, less radioactivity was incorporated in Calvin cycle intermediates and more in sucrose. carboxylations and photorespiration seemed to be inhibited. In the dark in both CO₂ conditions, ¹⁴C incorporation was found in malate and aspartate but also in serine and glycerate in high CO₂ conditions. In light these changes in metabolic pathways may be related with the deleterious effects recorded on leaves after long-term expositions to high partial pressure of CO₂.Abbreviations DHAP dihydroxyacetone phosphate - PEP phosphonenolpyruvate - PEPCK phosphonenolpyruvatecarboxykinase - PGA 3-phosphoglyceric acid - RUBPc ribulose 1,5-bisphosphate carboxylase     

A sensitive,simultaneous analysis of ribulose 1,5-bisphosphate carboxylase/oxygenase efficiencies: Graphical determination of the CO2/O2 specificity factor

A simple approach to determine CO₂/O₂ specificity factor () of ribulose 1,5-bisphosphate carboxylase/oxygenase is described. The assay measures the amount of CO₂ fixation at varying [CO₂]/[O₂] ratios after complete consumption of ribulose 1,5-bisphosphate (RuBP). Carbon dioxide fixation catalyzed by the carboxylase was monitored by directly measuring the moles of ¹⁴CO₂ incorporated into 3-phosphoglycerate (PGA). This measurement at different [CO₂]/[O₂] ratios is used to determine graphically by several different linear plots the total RuBP consumed by the two activities and the CO₂/O₂ specificity factor. The assay can be used to measure the amounts of products of the carboxylase and oxygenase reactions and to determine the concentration of the substrate RuBP converted to an endpoint amount of PGA and phosphoglycolate. The assay was found to be suitable for all [CO₂]/[O₂] ratios examined, ranging from 14 to 215 micromolar CO₂ (provided as 1–16 mM NaHCO₃) and 614 micromolar O₂ provided as 50% O₂. The procedure described is extremely rapid and sensitive. Specificity factors for enzymes of highly divergent values are in good agreement with previously published data.Abbreviations HEPPS N-(2-hydroxyethyl)piperazine-N-(3-propanesulfonic acid) - L large subunit of rubisco - PGA 3-phosphoglyceric acid - rubisco ribulose 1,5-bisphosphate carboxylase/oxygenase - RuBP d-ribulose 1,5-bisphosphate - S small subunit of rubisco - XuBP d-xylulose 1,5-bisphosphate     

Metabolic regulation of carbon flux during C4 photosynthesis

The potential for glycolate and glycine metabolism and the mechanism of refixation of photorespiratory CO₂ in leaves of C₄ plants were studied by parallel inhibitor experiments with thin leaf slices, different leaf cell types and isolated mitochondria of C₃ and C₄ Panicum species. CO₂ evolution by leaf slices of P. bisulcatum, a C₃ species, fed glycolate or glycine was light-independent and O₂-sensitive. The C₄ P. maximum and P. miliaceum leaf slices fed glycolate or glycine evolved CO₂ in the dark but not in the light. In C₄ species, dark CO₂ evolution was abolished by the addition of phosphoenolpyruvate (PEP)⁴. The addition of maleate, a PEP carboxylase inhibitor, resulted in photorespiratory CO₂ efflux by C₄ leaf slices in the light also. However, PEP and maleate had no effect on either glycolate-dependent O₂ uptake by the C₄ leaf slices or on glycolate and glycine metabolism in C₃ leaf slices. The rate of photorespiratory CO₂ evolution in the C₃ Panicum species was 3 times higher than that observed with the C₄ species. The ratio of glycolate-dependent CO₂ evolution to O₂ uptake in both groups was 1:2. Isolated C₄ mesophyll protoplasts or their mitochondria did not metabolize glycolate or glycine. However, both C₃ mesophyll protoplasts and C₄ bundle sheath strands readily metabolized glycolate and glycine in a light-independent, O₂-sensitive manner, and the addition of PEP or maleate had no effect. C₄ bundle sheath- and C₃-mitochondria were capable of oxidizing glycine. This oxidation was linked to the mitochondrial electron transport chain, was coupled to three phosphorylation sites and was sensitive to electron transport inhibitors. C₄ bundle sheath- and C₃-mitochondrial glycine decarboxylation was stimulated by oxaloacetate and NAD had no effect. In marked contrast, mitochondria isolated from C₄ mesophyll cells were incapable of oxidizing or decarboxylating added glycine. The results suggest that in leaves of C₄ plants bundle sheath cells are the primary site of O₂-sensitive photorespiratory CO₂ evolution and the PEP carboxylase present in the mesophyll cells has the Potential for efficiently refixing CO₂ before it escapes out of the leaf. The relative role of the PEP carboxylase mediated CO₂ pump and reassimilation of photorespiratory CO₂ are discussed in relation to the apparent lack of photorespiration in leaves of C₄ species.Abbreviations BSA bovine serum albumin - Chl chlorophyll - PEP phosphoenolpyruvate - Rbu-P ₂ ribulose 1,5-bisphosphate - Rib-5-P ribose-5-phosphate - Ru-5-P ribuluse-5-phosphate - FCCP carbonyl cyanide p-trifluoromethoxyphenylhydrazone Journal Series Paper, New Jersey Agricultural Experiment Station     

Gas-exchange of ears of cereals in response to carbon dioxide and light

Data for the maximum carboxylation velocity of ribulose-1,5-biosphosphate carboxylase, V_m, and the maximum rate of whole-chain electron transport, J_m, were calculated according to a photosynthesis model from the CO₂ response and the light response of CO₂ uptake measured on ears of wheat (Triticum aestivum L. cv. Arkas), oat (Avena sativa L. cv. Lorenz), and barley (Hordeum vulgare L. cv. Aramir). The ratio J_m/V_m is lower in glumes of oat and awns of barley than it is in the bracts of wheat and in the lemmas and paleae of oat and barley. Light-microscopy studies revealed, in glumes and lemmas of wheat and in the lemmas of oat and barley, a second type of photosynthesizing cell which, in analogy to the Kranz anatomy of C₄ plants, can be designated as a bundle-sheath cell. In wheat ears, the CO₂-compensation point (in the absence of dissimilative respiration) is between those that are typical for C₃ and C₄ plants.A model of the CO₂ uptake in C₃–C₄ intermediate plants proposed by Peisker (1986, Plant Cell Environ. 9, 627–635) is applied to recalculate the initial slopes of the A(p^c) curves (net photosynthesis rate versus intercellular partial pressure of CO₂) under the assumptions that the J_m/V_m ratio for all organs investigated equals the value found in glumes of oat and awns of barley, and that ribulose-1,5-bisphosphate carboxylase is redistributed from mesophyll to bundle-sheath cells. The results closely match the measured values. As a consequence, all bracts of wheat ears and the inner bracts of oat and barley ears are likely to represent a C₃–C₄ intermediate type, while glumes of oat and awns of barley represent the C₃ type.Abbreviations A net photosynthesis rate (mol·m^-2·s^-1) - J_m maximum rate of whole-chain electron transport (mol·e^-·m^-2·s^-1) - p^c (bar) intercellular partial pressure of CO₂ - PEP phosphoenolpyruvate - PPFD photosynthetic photon flux density (mol quanta·m^-2·s^-1) - RuBPCase ribulose bisphosphate carboxylase/oxygenase - RuBP ribulose bisphosphate - V_m maximum carboxylation velocity of RuBPCase (mol·m^-2·s^-1) - T^* CO₂ compensation point in the absence of dissimilative respiration (bar)     

The in-vivo response of the ribulose-1,5-bisphosphate carboxylase activation state and the pool sizes of photosynthetic metabolites to elevated CO2 inPhaseolus vulgaris L.

The short-term, in-vivo response to elevated CO₂ of ribulose-1,5-bisphosphate carboxylase (RuBPCase, EC 4.1.1.39) activity, and the pool sizes of ribulose 1,5-bisphosphate, 3-phosphoglyceric acid, triose phosphates, fructose 1,6-bisphosphate, glucose 6-phosphate and fructose 6-phosphate in bean were studied. Increasing CO₂ from an ambient partial pressure of 360–1600 bar induced a substantial deactivation of RuBPCase at both saturating and subsaturating photon flux densities. Activation of RuBPCase declined for 30 min following the CO₂ increase. However, the rate of photosynthesis re-equilibrated within 6 min of the switch to high CO₂, indicating that RuBPCase activity did not limit photosynthesis at high CO₂. Following a return to low CO₂, RuBPCase activation increased to control levels within 10 min. The photosynthetic rate fell immediately after the return to low CO₂, and then increased in parallel with the increase in RuBPCase activation to the initial rate observed prior to the CO₂ increase. This indicated that RuBPCase activity limited photosynthesis while RuBPCase activation increased. Metabolite pools were temporarily affected during the first 10 min after either a CO₂ increase or decrease. However, they returned to their original level as the change in the activation state of RuBPCase neared completion. This result indicates that one role for changes in the activation state of RuBPCase is to regulate the pool sizes of photosynthetic intermediates.Abbreviations and symbols A net CO₂ assimilation rate - C_a ambient CO₂ partial pressure - C_i intercellular CO₂ partial pressure - CABP 2-carboxyarabinitol 1,5-bisphosphate - k_cat catalytic turnover rate per RuBPCase molecule - PFD photon flux density (400 to 700 nm on an area basis) - PGA 3-phosphoglyceric acid - P_i orthophosphate - RuBP ribulose 1,5-bisphosphate - RuBPCase ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39)     

Photosynthetic carbon metabolism of the cool-temperate C4 grass Spartina anglica Hubb.

The aim of this work was to describe the photosynthetic carbon metabolism of the cooltemperate C₄ grass Spartina anglica. With the exception of pyruvate, phosphate dikinase and pyruvate kinase, the maximum catalytic activities in leaves of putative enzymes of the C₄ cycle of a phosphoenolpyruvate-carboxykinase C₄ plant were considerably in excess of the observed, steady-state rate of photosynthesis, and were comparable with the maximum catalytic activities of key enzymes of the reductive pentose-phosphate pathway. Radioactive carbon from ¹⁴CO₂ supplied to attached leaves during steady-state photosynthesis appeared first in malate and aspartate from which it moved to intermediates of the reductive pentose-phosphate pathway, and then to sucrose. These experiments show that photosynthetic carbon metabolism in this cool-temperate C₄ plant is similar to that of C₄ plants of hotter climates.     

On the mechanism of autotrophic fixation of CO2 bychloroflexus aurantiacus

The activity of two carboxylating enzymes was studied in the green filamentous bacteriumChloroflexus aurantiacus. The carboxylation reaction involving pyruvate synthase was optimized using¹⁴CO₂ and cell extracts. Pyruvate synthase was shown to be absent from cells ofCfl. aurantiacus OK-70 and present (in a quantity sufficient to account for autotrophic growth) in cells ofCfl. aurantiacus B-3. Differences in the levels of acetyl CoA carboxylase activity were revealed between cells of the strains studied grown under different conditions. The data obtained confirm the operation of different mechanisms of autotrophic CO₂ assimilation inCfl. aurantiacus B-3 andCfl. aurantiacus OK-70: in the former organism, it is the reductive cycle of dicarboxylic acids, and in the latter one, it is the 3-hydroxypropionate cycle.     

The capacity of phototrophic sulfur bacterium Thiocapsa roseopersicina for chemosynthesis

Purple sulfur bacterium Thiocapsa roseopersicina strain BBS requiring vitamin B₁₂ may grow in the dark in media containing no other organic compounds. Under such conditions the cells oxidize sulfide and thiosulfate with the use of O₂ and assimilate carbon dioxide. After 10–30 s assimilation of NaH¹⁴CO₃ about 60% of radioactivity is found in phosphorylated compounds characteristic for the reductive pentose phosphate cycle. The possibility of the function of this cycle in the dark in the presence of O₂ is confirmed by the capacity of cells grown under such conditions to synthesize ribulose-1,5-diphosphate carboxylase. All this evidence suggests the ability of T. roseopersicina to change from phototrophy to aerobic chemolithoautotrophy.