Photosynthetic carbon dioxide assimilation by Rhodospirillum rubrum

Summary Although Rhodospirillum rubrum, grown photoheterotrophically on malate, assimilates carbon dioxide less rapidly than it does when grown autotrophically, the difference is less marked than previously suggested.The rate of photoassimilation of carbon dioxide varies during batch culture on malate, reaching a maximum at about mid-exponential phase. It also varies with density and growth rate in a turbidostat continuous-flow culture on malate and increases with decreasing growth rate in a chemostat continuous-flow culture growing with limiting malate concentrations.The changing rates of carbon dioxide photoassimilation during photoheterotrophic growth under the various conditions are paralleled by changing activities of ribulose diphosphate carboxylase.Under conditions of maximum carbon dioxide fixation the rate by photoheterotrophic cultures approaches that shown by the bacterium growing autotrophically and is assimilated eight to ten times more slowly than is malate in chemostat cultures.The rate of carbon dioxide fixation also increases to that shown by autotrophic cells when photoheterotrophic cultures are deprived of malate, but without subjecting them to the conditions required for autotrophic growth.     

Light-dependent synthesis of glutamate inRhodospirillum rubrum

Summary The relative sizes of the macro amino acid pools inRhodospirillum rubrum, as measured by the incorporation of [¹⁴C]-carbon dioxide, were approximately the same in the four different cultures examined; mid-exponential phase and initial stationary phase batch culture organisms and two steady state turbidostat continuous cultures. Glutamate was the dominant amino acid labelled with lower levels of labelling in glutamine, aspartate, alanine and threonine to complete the major amino acid pools. Glutamine synthesis was a light-dependent process in the four cultures examined whereas the synthesis of glutamate was light-dependent only in the stationary phase batch culture organisms and in low cell density turbidostat organisms. These results are presented as physiological evidence for the activity of the ATP-requiring GS-GOGAT system for ammonia assimilation under certain growth conditions inRhodospirillum rubrum.     

VISCOUS POLYSACCHARIDE AND STARCH SYNTHESIS IN RHODELLA RETICULATA (PORPHYRIDIALES,RHODOPHYTA)1

Rhodella reticulata Deason, Butler and Rhyne produces copious amounts of a viscous polysaccharide (VP) during growth in batch cultures. The VPs accumulated on the cell surface and in the culture medium once cells ceased growth; starch concurrently accumulated within the cells. Light-saturated ¹⁴C-uptake declined steadily as the cells aged. Net synthesis rates for starch and mucilage were two- and four-fold lower, respectively, in non-growing cells than in growing cells, while the relative partitioning of newly-fixed carbon into these materials was not different. These data suggest that total photosynthetic loading, rather than partitioning into one specific pool, controls cellular synthesis rates. No preferential synthesis of VPs occurred during the stationary phase. The findings have important implications for the commercial production of VPs.     

INORGANIC CARBON TRANSPORT IN RELATION TO CULTURE AGE AND INORGANIC CARBON CONCENTRATION IN A HIGH-CALCIFYING STRAIN OF EMILIANIA HUXLEYI (PRYMNESIOPHYCEAE)1

The relationships among inorganic carbon transport, bicarbonate availability, intracellular pH, and culture age were investigated in high-calcifying cultures of Emiliania huxleyi (Lohmann) Hay & Mohler. Measurement of inorganic carbon transport by the silicone-oil centrifugation technique demonstrated that gadolinium, a potential Ca²⁺ channel inhibitor, blocked intracellular inorganic carbon uptake and photosynthetic ¹⁴CO₂₊ fixation in exponential-phase cells. In stationary-phase cells, the intracellular inorganic carbon concentration was unaffected by gadolinium. Gadolinium was also used to investigate the link between bicarbonate and Ca²⁺ transport in high-calcifying cells of E. huxleyi. Bicarbonate availability had significant and rapid effects on pH_i in exponential- but not in stationary-phase cells. 4′, 4′-Diisothiocyanostilbene-2,2′-disulfonic acid did not block bicarbonate uptake from the external medium by exponential-phase cells. Inorganic carbon utilization by exponential- and stationary-phase cells of Emiliania huxleyi was investigated using a pH drift technique in a closed system. Light-dependent alkalization of the medium by stationary-phase cells resulted in a final pH of 10.1 and was inhibited by dextran-bound sulphonamide, an inhibitor of external carbonic anhydrase. Exponential-phase cells did not generate a pH drift. Overall, the results suggest that for high-calcifying cultures of E. huxleyi the predominant pathway of inorganic carbon utilization differs in exponential and stationary phase cells of the same culture.     

CHLOROPHYLL,NITROGEN, AND PHOTOSYNTHETIC PATTERNS DURING GROWTH AND SENESCENCE OF TWO BLUE-GREEN ALGAE1,2

A standardized, multiflask, batch culture system was developed to study the processes of algal senescence in Anacystis nidulans and Phormidium molle Gom, var. tenuior W. et G. West. Growth data over a 3-year period gave reproducible and comparable time-course curves. Although A. nidulans is unicellular and P. molle filamentous, the patterns of change with age were similar. Mean logarithmic doubling times and carbon yields were, respectively, 6.9 hr and 390 mg C/liter for A. nidulans and 7.2 hr and 710 mg C/liter for P. molle. Chlorophyll concentration and photo-synthetic capacity per unit carbon rose rapidly during the logarithmic phase to maximum levels in either late log phase (P. molle) or early linear phase (A. nidulans), then fell throughout the declining growth phase to low levels in the stationary phase. Nitrate was rapidly exhausted from the medium during the period of logarithmic growth and stoichiometrically converted to particulate organic form; very little subsequent fixation of molecular nitrogen occurred. The phycocyanins were rapidly destroyed during the logarithmic phase while the carotenoids remained relatively constant throughout the whole growth period and then slowly declined. Preliminary electron micrographs showed a progressive deterioration in cellular ultrastructure, especially a reduction in the number of photosynthetic thylakoids, commenting in the linear growth phase. Analysis of the results suggests that occurrence of linear growth kinetics and termination of culture growth were caused by exhaustion of nitrate. The observed decreases in chlorophylls, phycocyanins, and photosynthetic capacity during active culture growth show that senescence effects may not be, as assumed, restricted to the stationary phase of growth.     

PHOSPHATE UPTAKE KINETICS IN EUGLENA GRACILIS (Z) (EUGLENOPHYCEAE) GROWN IN LIGHT/DARK CYCLES. II. PHASED PO4-LIMITED CULTURES1

The characteristics of phosphate uptake and photosynthetic capacity were studied in P-limited populations of Euglena gracilis Klebs (Z), using both P-limited batch cultures in stationary phase and cyclostat cultures grown on 14:10 LD. P uptake obeyed Michaelis-Menten kinetics between 0 and 150 μM PO₄ under both growth conditions. The value of V_max was 35% lower in the dark than in the light in the stationary phase cells. The value of K₈ was not affected by light conditions, and uptake was completely inhibited in the presence of 1 mm KCN. P uptake (at 2.0 μM PO₄) and photosynthetic capacity showed diel periodicity with peak rates occurring just before the beginning of the dark period for P uptake, and 8 h into the light period for photosynthetic capacity. V_max for P uptake increased by a factor of 1.5 over the light period, whereas K₈ remained constant at 1.4 μM PO₄. These patterns were displayed by both nondividing stationary phase cells and populations in which less than a third of the cells divided each day, indicating that the rhythmicity is not coupled to cell division.     

Control of photosynthetic carbon dioxide fixation by the boundary layer, stomata and ribulose 1,5-biphosphate carboxylase/oxygenase

Abstract Coefficients describing the sensitivity of the rate of photosynthetic carbon dioxide fixation to small changes in the stomatal conductance and boundary layer conductance are derived. These sensitivity or ‘control’ coefficients, together with those for the carboxylase and oxygenase activities of ribulose 1,5-bisphosphate carboxylase/oxygenase, are calculated from standard gas exchange data and apply under conditions where leaf temperature and water vapour concentration at the leaf surface remain largely constant. It is shown that the magnitude of the control coefficients depends on conditions such as photon flux density, ambient CO₂ concentration and relative humidity at the leaf surface. The extension of this analysis to encompass the sensitivity of the photosynthetic fluxes to changes in enzyme concentrations and kinetic properties is also discussed.     

Carbon dioxide fixation in Pyrobotrys stellata

The green alga Pyrobotrys stellata Korshik., an obligate phototroph, is unable to utilise carbon dioxide for growth, although assimilation of acetate is dependent on the photosynthetic process. The incorporation of ¹⁴CO₂ from ¹⁴C-bicarbonate into the cells of P. stellata is only 3% of that in Chlorella pyrenoidosa Chick. The activity of the key enzyme of the Calvin cycle, ribulose-1-5-diphosphate carboxylase, is very low in P. stellata, being only 7% of that in C. pyrenoidosa. The determination of the products of ¹⁴CO₂ fixation in intact cells confirms that ribulose-1-5-diphosphate activity is very low in P. stellata, since little carbon-14 is found in 1–3 diphosphoglyceric acid, the product of carboxylation of ribulose-1-5-diphosphate. It is concluded that the inability of P. stellata to utilize carbon dioxide for growth in the light is probably the result of the low ribulose-1-5-diphosphate carboxylase activity in the organism.     

Photoautotrophic growth of cell suspension cultures fromChenopodium rubrum in an airlift fermenter

Summary This communication describes the construction and operation of an airlift fermenter for the photoautotrophic growth of cell suspension cultures fromChenopodium rubrum. The basic batch culture unit provides a culture of 1.51 volume, sufficient to permit frequent aseptic sampling. It can be maintained at any desired temperature and aerated to different extents. Using an initial cell density of about 400,000 cells per ml suspension, the increase in cell number is 270% after a 14 days' growth period, although the stationary phase of growth is not yet reached. The transfer of photoautotrophic cell suspensions fromChenopodium rubrum from stationary growth into the large volume of fresh culture medium in the airlift fermenter results in an immediate protein formation, followed by an exponential phase of cell division, whereas rapid chlorophyll accumulation is delayed by 2 days.The growth capacities of photoautotrophic fermenter cultures including protein and chlorophyll formation as well asin vitro activities of the ribulosebisphosphate carboxylase and the phosphoenolpyruvate carboxylase are greatly lower as compared to photoautotrophic cells propagated in standard two-tier culture vessels using 30 ml culture medium. However the pattern of change in the activities of carboxylation enzymes is quite similar in both culture systems.Photoautotrophic cell suspensions fromChenopodium rubrum grown in an airlift fermenter assimilate about 90 mol CO₂/mg chlorophyll × hour. Dark CO₂ fixation is about 1.5% of the light values.Abbreviations PEF phosphoenolpyruvate - RuDP ribulosebisphosPhate - NS ground glass joints of standardized size made from Duran glass, Schott, Germany     

Effects of nitrogen starvation on the photosynthetic physiology of a tropical marine microalga Rhodomonas sp. (Cryptophyceae)

The effects of nitrogen starvation on biomass composition and photosynthetic function were examined in the marine cryptophyte Rhodomonas sp. Batch-cultured cells in N-sufficient medium showed a 2.5-fold increase in total carbohydrate content, and a 33% increase in cell volume when the cultures reached the stationary growth phase. These cultures also increased the ratio of phycoerythrin (PE)/hydrosoluble proteins from 6 to 22% by the 4th and 10th day of culture, respectively. In contrast, light-saturated photosynthetic activity (P_m) progressively decreased, and the value obtained at the beginning of the stationary phase was about 45% of that obtained for cells in the late exponential growth phase. Transfer to N-lacking medium caused a 3.2-fold increase in cell volume. N starvation also triggered a rapid decline in N-containing compounds such as hydrosoluble proteins and photosynthetic pigments, causing an almost complete loss of PE. The ratio of PE/hydrosoluble proteins decreased from 6 to 1% after 6 d of N deprivation. Furthermore, the PSII fluorescence capacity declined under N-starved conditions, which caused a pronounced decrease in both the P_m (circa 90%) and the apparent photosynthetic efficiency (circa 55%). Under these conditions, photosynthetically fixed carbon was used to synthesize large amounts of carbohydrates. We suggest that, in addition to the role of phycoerythrin as a light-harvesting pigment, Rhodomonas sp. responds to N-depleted conditions by mobilizing combined nitrogen from biliproteins.     

Changes in Membrane Fatty Acid Composition and Δ12-Desaturase Activity during Growth of Acanthamoeba castellanii in Batch Culture

ABSTRACT. Growth of Acanthamoeba castellanii in batch culture at 30° C was associated with marked changes in cellular fatty acid composition. The largest change occurred in the linoleate to oleate ratio, which was maximal in early- to mid-exponential phase cultures but decreased approximately 10-fold as cells approached stationary phase. The higher degree of lipid unsaturation in young cultures was accentuated by a greater proportion of 20-carbon polyunsaturated fatty acids than in stationary phase cultures. The unsaturation index (average number of double bonds per fatty acid) was maximal in mid-exponential phase cultures after 24 hours growth. Incorporation of [1-¹⁴C]acetate into polyunsaturated fatty acids in short-term (2 hour) experiments was high in 12 and 24 hour old cultures, where linoleate and eicosadienoate accounted for up to 26% of total labelled fatty acids. Incorporation of [1-¹⁴CJacetate into these fatty acids was negligible in stationary phase cultures. These results were correlated with changes in the specific activity of the Δ12-desaturase. Δ12-Desaturase activity was greatest in microsomal membranes isolated from early- to mid-exponential phase cells, but declined by approximately 50% as cultures progressed towards stationary phase. Membrane fractionation studies revealed that although some differences in fatty acid composition between plasma-membrane, mitochondrial (enriched), and microsomal membrane fractions were evident, the large changes in lipid unsaturation in whole cells of A. castellanii could not be accounted for by differential development of particular subcellular membranes.     

Relations between carbon dioxide fluxes and environmental factors of Kobresia humilis meadows and Potentilla fruticosa meadows

Carbon dioxide fluxes of Kobresia humilis and Potentilla fruticosa shrub meadows, two typical ecosystems in the Qinghai-Tibet Plateau, were measured by eddy covariance technology and the data collected in August 2003 were employed to analyze the relations between carbon dioxide fluxes and environmental factors of the ecosystems. August is the time when the two ecosystems reach their peak leaf area indexes and stay stable, and also the period when the net carbon absorptions of Kobresia humilis and Potentilla fruticosa shrub meadows reach 56.2 g C·m⁻² and 32.6 g C·m⁻², with their highest daily carbon dioxide absorptions standing at 12.7 μmol·m⁻²·s⁻¹ and 9.3 μmol·m⁻²·s⁻¹, and their highest carbon discharges at 5.1 μmol·m⁻²·s⁻¹ and 5.7 μmol·m⁻²·s⁻¹, respectively. At the same photosynthetic photo flux densities (PPFD), the carbon dioxide-uptake rate of the Kobresia humilis meadow is higher than that of the Potentilla fruticosa shrub meadow; where the PPFD are higher than 1,200 μmol·m⁻²·s⁻¹. The carbon dioxide uptake rates of the two ecosystems declined as air temperature increased, but the carbon dioxide uptake rate of the Kobresia humilis meadow decreased more quickly (−0.086) than that of the Potentilla fruticosa shrub meadow (−0.016). Soil moistures exert influence on the soil respirations and this varies with the vegetation type. The daily carbon dioxide absorptions of the ecosystems increase with increased diurnal temperature differences and higher diurnal temperature differences result in higher carbon dioxide exchanges. There exists a negative correlation between the vegetation albedos and the carbon dioxide fluxes. Translated from Acta Bot Boreal—Occident Sin, 2006, 26(1): 133–142 [译自: 西北植物学报]     

Growth and activities of enzymes of primary metabolism in batch cultures of Catharanthus roseus cell suspension under different pCO2 conditions

In vitro enzyme activities of glycolysis, pentose-phosphate pathway and dark CO₂ fixation were assayed in batch cultures of heterotrophic Catharanthus roseus cells under various gassing rates and partial pressures of carbon dioxide. Detrimental effects of low pCO₂ culture conditions on the growth characteristics could be linked to marked changes in levels of enzymes of primary metabolism during growth. The enzyme levels observed during the early stages of growth were found to be more stable when a constant pCO₂ (20 mbar) was maintained and enabled exponential growth to be reached more rapidly.The importance of carbon dioxide as a conditioning factor of the culture medium is discussed.     

Activity of the Enzymes of Carbon Metabolism in Sulfobacillus sibiricus under Various Conditions of Cultivation

The thermoacidophilic iron-oxidizing chemolithotroph Sulfobacillus sibiricus N1^T is characterized by steady growth and amplified cell yield when grown in vigorously aerated medium containing Fe²⁺, glucose, and yeast extract as energy sources. In this case, carbon dioxide, glucose, and yeast extract are used as carbon sources. Glucose is assimilated through the fructose-bisphosphate pathway and the pentose-phosphate pathway. The glyoxylate bypass does not function in S. sibiricus, and the tricarboxylic acid cycle is disrupted at the level of 2-oxoglutarate dehydrogenase. The presence of ribulose-bisphosphate carboxylase indicates that carbon dioxide fixation proceeds through the Calvin cycle. The activity of ribulose-bisphosphate carboxylase is highest in autotrophically grown cells. The cells also contain pyruvate carboxylase, phosphoenolpyruvate carboxylase, phosphoenolpyruvate carboxykinase, and phosphoenolpyruvate carboxytransphosphorylase.     

Microbial metabolism of fluoranthene: isolation and identification of ring fission products

Summary The degradation of fluoranthene by pure cultures of Alcaligenes denitrificanss WW1, isolated from contaminated soil samples, was investigated. The strain showed maximum degradation rates of 0.3 mg fluoranthene/ml per day. A denitrificans was able to utilize also naphthalene, 1- and 2-methylnaphthalene, phenanthrene, and anthracene as sole carbon sources and to co-metabolize fuuorence, pyrene, and benzo(a)anthracene. During growth on fluoranthene in batch culture two metabolic products that were completely degraded before growth entered the stationary phase were detected in the culture fluid. Anslyses by UV, mass and NMR spectroscopy identified the products as acenaphthenone and 3-hydroxymethyl-4,5-benzocoumarine. Fluoranthene-grown resting cells of A. denitrificans showed degradative activity towards 2,3-dihydroxybenzoic acid, pyrogallol, salicylic acid, and catechol. The enzymatic activities in extracts of fluoranthene-induced cells indicate a meta ring fission involved in the degradation of fluoranthene. From these data new aspects of the biodegradative pathway of fluoranthene have been predicted.     

Mixotrophic growth of Thiobacillus A2 on acetate and thiosulfate as growth limiting substrates in the chemostat

During heterotrophic growth on acetate, in batch culture, the autotrophic growth potential of Thiobacillus A2, i.e. the capacity to oxidize thiosulfate and to fix carbon dioxide via the Calvin cycle, was completely repressed. The presence of thiosulfate in a batch culture with acetate as the organic substrate partly released the repression of the thiosulfate oxidizing system. Cultivation of the organism in continuous culture at a dilution rate of 0.05 h^-1 with different concentration ratios of thiosulfate and acetate in the reservoir medium led to mixotrophic growth under dual substrate limitation. Growth on the different mixtures of acetate and thiosulfate yielded upto 30% more cell dry weight than predicted from the growth yields on comparable amounts of these substrates separately. The extent to which the carbon dioxide fixation capacity and the maximum thiosulfate and acetate oxidation capacity are repressed appeared to be a function of the thiosulfate to acetate concentration ratio in the reservoir medium. The results of ¹⁴C-acetate assimilation experiments and of gas-analysis demonstrated that the extent to which acetate was assimilated depended also on the substrate ratio in the inflowing medium. Under the different growth conditions surprisingly little variation was found in some tri-carboxylic acid cycle enzyme activities. Cultivation of T. A2 at different growth rates with a fixed mixture of thiosulfate (18 mM) and acetate (11 mM) in the medium, showed that dual substrate limitation occured at dilution rates ranging from 0.03–0.20 h^-1.Abbreviations PPO 2,5-diphenoloxazol - RubPCase Ribulose-1,5-bisphophate carboxylase - Tris tris (hydroxymethyl) aminomethane - EDTA ethylenediaminetetra-acetic acid     

Reversible inhibition of CO2 fixation by ribulose 1,5‐bisphosphate carboxylase/oxygenase through the synergic effect of arsenite and a monothiol

The activity of the photosynthetic carbon‐fixing enzyme, ribulose 1,5‐bisphosphate carboxylase/oxygenase (Rubisco), is partially inhibited by arsenite in the millimolar concentration range. However, micromolar arsenite can fully inhibit Rubisco in the presence of a potentiating monothiol such as cysteine, cysteamine, 2‐mercaptoethanol or N‐acetylcysteine, but not glutathione. Arsenite reacts specifically with the vicinal Cys172‐Cys192 from the large subunit of Rubisco and with the monothiol to establish a ternary complex, which is suggested to be a trithioarsenical. The stability of the complex is strongly dependent on the nature of the monothiol. Enzyme activity is fully recovered through the disassembly of the complex after eliminating arsenite and/or the thiol from the medium. The synergic combination of arsenite and a monothiol acts also in vivo stopping carbon dioxide fixation in illuminated cultures of Chlamydomonas reinhardtii. Again, this effect may be reverted by washing the cells. However, in vivo inhibition does not result from the blocking of Rubisco since mutant strains carrying Rubiscos with Cys172 and/or Cys192 substitutions (which are insensitive to arsenite in vitro) are also arrested. This suggests the existence of a specific sensor controlling carbon fixation that is even more sensitive than Rubisco to the arsenite–thiol synergism.     

Effects of Cytokinin Preparations on the Stability of the Photosynthetic Apparatus of Two Wheat Cultivars Experiencing Water Deficiency

Carboxylase activity of the key enzyme of carbon metabolism, ribulose-bisphosphate carboxylase/oxygenase (RuBisCO; EC 4.1.1.39), and phosphoenolpyruvate carboxylase (PEPC; EC 4.1.1.31), as well as the intensity of carbon dioxide photosynthetic assimilation in young seedlings and adult leaves of the wheat Triticum aestivum L. cultivars Mironovskaya 808 (a more tolerant) and Lyutestsens758 (a less tolerant), were compared under conditions of progressive water deficiency. The water stress had more pronounced negative effects on all the studied characteristics of the photosynthetic apparatus of the cultivar Lyutestsens758. Its seedlings were more sensitive to water stress. Compounds with a cytokinin activity (6-benzylaminopurine, thidiazuron, kartolin 2, and kartolin 4) played a protective role, increasing the stability of the photosynthetic apparatus under conditions of water deficiency. Preparations of kartolins displayed the maximum protective effect.     

Growth characteristics of hormone and vitamin independent photoautotrophic cell suspension cultures fromChenopodium rubrum

Summary This communication reports the photoautotrophic growth of hormone and vitamin independent cell suspension cultures ofChenopodium rubrum. The transfer of cells from stationary growth into fresh culture medium results in a high protein formation, followed by an exponential phase of cell division, whereas the onset of rapid chlorophyll formation is delayed for 4 days. At the stage of most rapid cell division there is no net synthesis of starch and sugar. When the cells enter stationary growth, there is a progressive accumulation of chlorophyll, sugar, and starch.Photoautotrophic cell cultures assimilate about 80–90 mol CO₂/mg chlorophyll X hour. Dark CO₂ fixation is about 3.7% to 2.2% of the light values during exponential and stationary growth, respectively. As shown by short-term¹⁴CO₂ fixation, CO₂ is predominantly assimilated through ribulosebisphosphate carboxylase via the Calvin pathway. There is a significant increase in the¹⁴C label of C₄ carboxylic acids in exponentially dividing cells as compared to cells from stationary growth. Thein vitro activity of phosphoenolpyruvate carboxylase and ribulosebisphosphate carboxylase is almost equal during exponential cell division. A decrease in cell division activity is accompanied by a significant change in the specific activities of both carboxylation enzymes. In non dividing cells from stationary growth the activity of ribulosebisphosphate carboxylase is greately enhanced and that of phosphoenolpyruvate carboxylase is reduced, documenting the development of carboxylation capacities typical for C₃-plants.The experimental results provide evidence that phosphoenolpyruvate carboxylase activity might be regulated by ammonia and could be involved in anaplerotic CO₂ fixation which supplies carbon skeletons of the citric acid cycle.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - EDTA ethylene-diamine-tetraacetic acid - FDP fructose bisphosphate - F-6-P fructose-6-phosphate - G-6-P glucose-6-phosphate - HEPES N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid - PGA 3-phosphoglyceric acid - PEP phosphoenolpyruvate - RuDP ribulosebisphosphate