Separation of phosphate esters and algal extracts by thin-layer electrophoresis and chromatography
About: This article is published in Journal of Chromatography A.The article was published on 1969-02-25. It has received 94 citations till now.
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TL;DR: Starch synthesis in leaves was increased by phosphate starvation or by treatments which decreased cytoplasmic orthophosphate levels (such as mannose feeding) and it is suggested that this control is exerted via the phosphate translocator and the known allosteric regulation of ADP-glucose pyrophosphorylase.
Abstract: Starch synthesis in leaves was increased by phosphate starvation or by treatments which decreased cytoplasmic orthophosphate levels (such as mannose feeding). Usually less than 30% of the total carbon fixed during CO(2) assimilation was incorporated into starch in spinach (Spinacia oleracea L.), spinach beet (Beta vulgaris), and tobacco (Nicotiana tabacum) leaves.In isolated spinach chloroplasts, formation of starch from CO(2) was usually less than in leaves. In the absence of significant levels of 3-phosphoglycerate, concentrations of phosphate as low as 1 mm (in the medium) or 10 mm (in the stroma) almost completely inhibited starch synthesis. The inhibitory action of phosphate could be overcome by 3-phosphoglycerate. The controlling factor of starch synthesis appeared to be the ratio of phosphoglycerate to orthophosphate rather than the stromal hexose monophosphate concentration, and it is suggested that this control is exerted via the phosphate translocator and the known allosteric regulation of ADP-glucose pyrophosphorylase. Starch synthesis was also favored by the presence of dihydroxyacetone phosphate and by high light and high temperature. Oxygen was inhibitory, probably owing to carbon drain into glycolate. Starch formation by intact chloroplasts could not be promoted by added glucose or glucose 6-phosphate.Starch mobilization in the dark was promoted by orthophosphate and phosphate-dependent mobilization was inhibited by phosphoglycerate. The principal products of starch breakdown in the presence of phosphate were the transport metabolites dihydroxyacetone phosphate and 3-phosphoglycerate. Formation of these compounds from starch was stimulated by ATP or oxaloacetate. In a phosphate-independent reaction, starch was also converted to neutral products such as maltose and glucose. The rates of phosphate-dependent starch degradation phosphorolysis were very much higher than those of starch hydrolysis for which there was no phosphate requirement.
374 citations
TL;DR: It is shown that intact chloroplasts are able to fix CO2 in the dark, yielding 3-phosphoglycerate, and demonstrates that CO2 fixation in thedark is not possible, unless the pH in the medium is artificially raised to pH 8.8.
352 citations
TL;DR: The results show Bienertia utilizes strict compartmentation of organelles and enzymes within a single cell to effectively mimic the spatial separation of Kranz anatomy, allowing it to function as a C4 plant having suppressed photorespiration; this raises interesting questions about evolution of C4 mechanisms.
Abstract: Summary Kranz anatomy, with its separation of elements of the C 4 pathway between two cells, has been an accepted criterion for function of C4 photosynthesis in terrestrial plants. However, Bienertia cycloptera (Chenopodiaceae), which grows in salty depressions of Central Asian semi-deserts, has unusual chlorenchyma, lacks Kranz anatomy, but has photosynthetic features of C 4 plants. Its photosynthetic response to varying CO2 and O2 is typical of C4 plants having Kranz anatomy. Lack of night-time CO2 fixation indicates it is not acquiring carbon by Crassulacean acid metabolism. This species exhibits an independent, novel solution to function of the C4 mechanism through spatial compartmentation of dimorphic chloroplasts, other organelles and photosynthetic enzymes in distinct positions within a single chlorenchyma cell. The chlorenchyma cells have a large, spherical central cytoplasmic compartment interconnected by cytoplasmic channels through the vacuole to the peripheral cytoplasm. This compartment is filled with mitochondria and granal chloroplasts, while the peripheral cytoplasm apparently lacks mitochondria and has grana-deficient chloroplasts. Immunolocalization studies show enzymes compartmentalized selectively in the CC compartment, including Rubisco in chloroplasts, and NAD-malic enzyme and glycine decarboxylase in mitochondria, whereas pyruvate, Pi dikinase of the C 4 cycle is localized selectively in peripheral chloroplasts. Phosphoenolpyruvate carboxylase, a cytosolic C4 cycle enzyme, is enriched in the peripheral cytoplasm. Our results show Bienertia utilizes strict compartmentation of organelles and enzymes within a single cell to effectively mimic the spatial separation of Kranz anatomy, allowing it to function as a C 4 plant having suppressed photorespiration; this raises interesting questions about evolution of C 4 mechanisms.
163 citations
Cites methods from "Separation of phosphate esters and ..."
...Separation and identi®cation of the labelled photosynthetic products were accomplished using two-dimensional thin-layer electrophoresis and chromatography methods (Moore and Seeman, 1990; Schurmann, 1969)....
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TL;DR: The mechanism of CO2 fixation in autotrophically grown cells was studied in this article, and the presence of ribulose-1,5-bis-phosphate carboxylase and phosphoribulokinase could not be demonstrated.
Abstract: 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 (δ13C) 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 α-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.
132 citations
TL;DR: 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, indicating a co-function of the C3 and C4 cycles in CO2 assimilation.
Abstract: 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.
113 citations
References
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TL;DR: In this article, Feue r Iangen et al. discuss the effects of farblose's farbloses on the formation of the Assozians in the Hebrew Bible.
Abstract: Die A n a l y s e n r e s u l t a t e s t i m m e n m i t den b e r e c h n e t e n W e r t e n iiberein. Die I n d i u m u n d Ga l l iumalky le s ind farblose, l i ch tb reehende Flf iss igkei ten, die ~ul3erst luff u n d feueht igke i t sempf ind l i ch s ind u n d sich in o rgan i schen L6sungs \" m i t t e l n wie .~ther, T e t r a h y d r o f u r a n , Benzol, Chloroform, Petrol&ther le icht 16sen. Ledigl ich Ind iumt r i i so -p ropy l k o n n t e b i she r noch n i ch t farblos, sonde rn s t e t s n u r als gelbe Flfissigkei t e rha l t en werden . W e i t e r h i n s ind die S u b s t a n z e n l ichtempf indl ich . Die I n d i u m a l k y l e s ind sowohl in t h e r m i s c h e r H i n s i c h t als a u c h gegenfiber Sauers to f f u n d :Feucht igke i t ins tab i le r als die e n t s p r e c h e n d e n Gal l iuinalkyle . So s ind sie alle an der L u f t se lbs tentzf indl ich , w~hrend z .B. Gal l iumt r i -n -p ropy l u n d h6he re Ga l l iumalky le an der L u f t n i ch t mel~r Feue r Iangen , s ich aber noch s t f i rmisch u n t e r s t a rke r R a u c h en tw ick lung zersetzen. M o l e k u l a r g e w i c h t s u n t e r s u c h u n g e n in Benzol d e u t e n je n a c h V e r b i n d u n g u n d molare r K o n z e n t r a t i o n auf Assoz ia t ion h i m Die Assoz ia t ionsgrade bewegen s ich in den Grenzen I b i s 1,7. Die U n t e r s u c h u n g e n we rden for tgese tz t .
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