Glycolytic activity in embryos of Pisum sativum and of non-dormant or dormant seeds of Avena sativa L. expressed through activities of PFK and PPi-PFK

Summary A quantative cytochemical assay for PP_i-PFK activity in the presence of Fru-2,6-P₂ is described along with its application to determine levels of activity in embryos of Pisum sativum and Avena sativa. The activity of ATP-PFK has also been studied in parallel as have PFK activities during the switch from dormant to non-dormant embryos in Avena sativa. PP_i-PFK activity, has been demonstrated in all tissues of Pisum sativum embryos and of Avena sativa embryos including the scutellum and the aleurone layers. The PP_i-PFK activity was greater than that of ATP-PFK in both dormant and non-dormant seeds though with only marginally more activity in the dormant as opposed to the non-dormant state.Abbreviations AMP adenosine monophosphate - ATP adenosine triphosphate - Fru-1,6-P₂ fructose 1,6-bisphosphate - Fru-2,6-P₂ fructose 2,6-bisphosphate - Fru-6-P fructose 6-phosphate - FB Pase 2 fructose 2,6-bisphosphatase (EC 3.1.3.46) - Gl-3-PD glyceraldehyde-3-phosphate dehydrogenase - NAD nicotinamide adenine dinucleotide - NBT nitroblue tetrazolium - PEP phosphoenolpyruvate - PFK 6-phosphofructokinase (EC 2.7.1.11) - PFK₂ 6-phosphofructo-2-kinase (EC 2.7.1.105) - PP_i pyrophosphate - PP_i-PFK pyrophosphate: fructose 6-phosphate 1-phosphotransferase (EC 2.7.1.90) - PVA polyvinyl alcohol (G04/140 Wacke Chemical Company)     

The role of inorganic phosphate in the regulation of C4 photosynthesis

Inorganic phosphate participates in many fundamental processes within the plant cell. Its broad influence on plant metabolism is related to such key operations as metabolite transport, enzyme regulation and carbohydrate metabolism in general. This review discusses these topics with special emphasis on the role assigned to this ubiquitous anion within the C₄ pathway of photosynthesis.Abbreviations DHAP dihydroxyacetone phosphate - Ga3P glyceraldehyde-3-phosphate - NAD(P)-ME-NAD(P) dependent malic enzyme - PEP phosphoenolpyruvate - 3-PGA 3-phosphoglycerate - PFK and PFP-ATP- and PPi dependent fructose-6-phosphate 1-phosphotransferase - PPDK pyruvate:orthophosphate dikinase - RPPC reductive pentose-phosphate cycle - RuBisCO ribulose bisphosphate carboxylase-oxygenase - SPS sucrose-6-phosphate synthase     

Mechanisms of glycolytic control during hibernation in the ground squirrel Spermophilus lateralis

Summary The mechanisms of glycolytic rate control during hibernation in the ground squirrel Spermophilus lateralis were investigated in four tissues: heart, liver, kidney, and leg muscle. Overall glycogen phosphorylase activity decreased significantly in liver and kidney to give 50% or 75% of the activity found in the corresponding euthermic organs, respectively. The concentration of fructose-2,6-bisphosphate (F-2,6-P₂) decreased significantly in heart and leg muscle during hibernation to 50% and 80% of euthermic tissue concentrations, respectively, but remained constant in liver and kidney. The overall activity of pyruvate dehydrogenase (PDH) in heart and kidney from hibernators was only 4% of the corresponding euthermic values. Measurements of phosphofructokinase (PFK) and pyruvate kinase (PK) kinetic parameters in euthermic and hibernating animals showed that heart and skeletal muscle had typical rabbit skeletal M-type PFK and M₁-type PK. Liver and kidney PFK were similar to the L-type enzyme from rabbit liver, whereas liver and kidney PK were similar to the M₂ isozyme found primarily in rabbit kidney. The kinetic parameters of PFK and PK from euthermic vs hibernating animals were not statistically different. These data indicate that tissue-specific phosphorylation of glycogen phosphorylase and PDH, as well as changes in the concentration of F-2,6-P₂ may be part of a general mechanism to coordinate glycolytic rate reduction in hibernating S. lateralis.Abbreviations ADP adenosine diphosphate - AMP adenosine monophosphate - ATP adenonine triphoshate - EDTA ethylenediaminetetra-acetic acid - EGTA ethylene glycol tetra-acetic acid - F-6-P fructose 6-phosphate - F-1,6-P₂ fructose 1,6-bisphosphate - F-2,6-P₂ fructose-2,6-bisphosphate - K _a activation coefficient - I₅₀ concentration of inhibitor which reduces control activity by 50% - PDH pyruvate dehydrogenase - PEP phosphoenolpyruvate - PFK 6-phosphofructo-1-kinase - PK pyruvate kinase     

Molecular biology of C4 phosphoenolpyruvate carboxylase: Structure,regulation and genetic engineering

Three to four families of nuclear genes encode different isoforms of phosphoenolpyruvate (PEP) carboxylase (PEPC): C₄-specific, C₃ or etiolated, CAM and root forms. C₄ leaf PEPC is encoded by a single gene (ppc) in sorghum and maize, but multiple genes in the C₄-dicot Flaveria trinervia. Selective expression of ppc in only C₄-mesophyll cells is proposed to be due to nuclear factors, DNA methylation and a distinct gene promoter. Deduced amino acid sequences of C₄-PEPC pinpoint the phosphorylatable serine near the N-terminus, C₄-specific valine and serine residues near the C-terminus, conserved cysteine, lysine and histidine residues and PEP binding/catalytic sites. During the PEPC reaction, PEP and bicarbonate are first converted into carboxyphosphate and the enolate of pyruvate. Carboxyphosphate decomposes within the active site into Pi and CO₂, the latter combining with the enolate to form oxalacetate. Besides carboxylation, PEPC catalyzes a HCO₃ ^--dependent hydrolysis of PEP to yield pyruvate and Pi. Post-translational regulation of PEPC occurs by a phosphorylation/dephosphorylation cascade in vivo and by reversible enzyme oligomerization in vitro. The interrelation between phosphorylation and oligomerization of the enzyme is not clear. PEPC-protein kinase (PEPC-PK), the enzyme responsible for phosphorylation of PEPC, has been studied extensively while only limited information is available on the protein phosphatase 2A capable of dephosphorylating PEPC. The C₄ ppc was cloned and expressed in Escherichia coli as well as tobacco. The transformed E. coli produced a functional/phosphorylatable C₄ PEPC and the transgenic tobacco plants expressed both C₃ and C₄ isoforms. Site-directed mutagenesis of ppc indicates the importance of His¹³⁸, His⁵⁷⁹ and Arg⁵⁸⁷ in catalysis and/or substrate-binding by the E. coli enzyme, Ser⁸ in the regulation of sorghum PEPC. Important areas for further research on C₄ PEPC are: mechanism of transduction of light signal during photoactivation of PEPC-PK and PEPC in leaves, extensive use of site-directed mutagenesis to precisely identify other key amino acid residues, changes in quarternary structure of PEPC in vivo, a high-resolution crystal structure, and hormonal regulation of PEPC expression.Abbreviations OAA oxalacetate - PEP phosphoenolpyruvate - PEPC PEP carboxylase - PEPC-PK PEPC-protein kinase - PPDK pyruvate, orthophosphate dikinase - Rubisco ribulose 1,5-bis-phosphate carboxylase/oxygenase - CAM Crassulacean acid metabolism     

Accumulation of fructose 1,6-bisphosphate in mutant cells of mucoid Pseudomonas aeruginosa as an evidence of phosphofructokinase activity

Phosphoglucose isomerase negative mutant of mucoid Pseudomonas aeruginosa accumulated relatively higher concentration of fructose 1,6-bisphosphate (Fru-1,6-P₂) when mannitol induced cells were incubated with this sugar alcohol. Also the toluene-treated cells of fructose 1,6-bisphosphate aldolase negative mutant of this organism produced Fru-1,6-P₂ from fructose 6-phosphate in presence of ATP, but not from 6-phosphogluconate. The results together suggested the presence of an ATP-dependent fructose 6-phosphate kinase (EC 2.7.1.11) in mucoid P. aeruginosa.Abbreviations ALD Fru-1,6-P₂ aldolse - DHAP dihydroxyacetone phosphate - F6P fructose 6-phosphate - G6P glucose 6-phosphate - Gly3P glyceraldehyde 3-phosphate - KDPG 2-keto 3-deoxy 6-phosphogluconate - PFK fructose 6-phosphate kinase - PGI phosphoglucose isomerase - 6PG 6-phosphogluconate     

Screening of certain mangroves for photosynthetic carbon metabolic pathway

The mangroves Rhizophora lamarkii, Ceriops roxburghiana, Bruguiera gymnorrhiza, Aegiceras corniculatum, and Lumnitzera racemosa were screened for their carbon metabolic pathways by measuring net photosynthetic rate (P _N), ¹³C discrimination rate, leaf anatomy, titratable acidity, and activities of phosphoenolpyruvate carboxylase, NADH-malate dehydrogenase, alanine aminotransferase, aspartate aminotransferase, and pyruvate phosphate dikinase. The tested mangroves had a well developed succulence, opening of stomata during day time and closure in the night hours, and absence of diurnal fluctuation of organic acids in their leaves which excludes the possibility of these species being CAM plants. Moreover, the leaf anatomy had not exhibited Kranz syndrome. The high values of discrimination against ¹³C, low P _N, high CO₂ compensation concentration, and the activities of aminotransferases in the direction of alanine formation suggest that the species may follow C₃ mode of carbon metabolic pathway.     

Role of fructose-1,6-bisphosphatase, fructose phosphotransferase, and phosphofructokinase in saccharide metabolism of four C3 grassland species under elevated CO2

We studied the effects of 15-months of elevated (700 μmol mol⁻¹) CO₂ concentration (EC) on the CO₂ assimilation rate, saccharide content, and the activity of key enzymes in the regulation of saccharide metabolism (glycolysis and gluconeogenesis) of four C₃ perennial temperate grassland species, the dicots Filipendula vulgaris and Salvia nemorosa and the monocots Festuca rupicola and Dactylis glomerata. The acclimation of photosynthesis to EC was downward in F. rupicola and D. glomerata whereas it was upward in F. vulgaris and S. nemorosa. At EC, F. rupicola and F. vulgaris leaves accumulated starch while soluble sugar contents were higher in F. vulgaris and D. glomerata. EC decreased pyrophosphate-D-fructose-6-phosphate l-phosphotransferase (PFP, EC 2.7.1.90) activity assayed with Fru-2,6-P₂ in F. vulgaris and D. glomerata and increased it in F. rupicola and S. nemorosa. Growth in EC decreased phosphofructokinase (PFK, EC 2.7.1.11) activity in all four species, the decrease being smallest in S. nemorosa and greatest in F. rupicola. With Fru-2,6-P2 in the assay medium, EC increased the PFP/PFK ratio, except in F. vulgaris. Cytosolic fructose-1,6-bisphosphatase (Fru-1,6-P₂ase, EC 3.1.3.11) was inhibited by EC, the effect being greatest in F. vulgaris and smallest in F. rupicola. Glucose-6-phosphate dehydrogenase (G6PDH EC 1.1.1.49) activity was decreased by growth EC in the four species. Activity ratios of Fru-1,6-P₂ase to PFP and PFK suggest that EC may shift sugar metabolism towards glycolysis in the dicots.     

Kinetic studies of pyruvate kinase duringin vitro differentiation of GM-CFC haemopoietic precursor and bone marrow cells in mice

Pyruvate kinase studies in the granulocyte-macrophage lineage duringin vitro differentiation have been performed using culture techniques on GM-CFC cells and a study has also been done in bone marrow cells.The enzyme exhibits biphasic behaviour with respect to both of its substrates in cells derived fromin vitro cultures at 5 and 7 days of incubation period. However in bone marrow cells these kinetics are only observed for ADP.The different kinetic behaviour of pyruvate kinase toward Fru-1,6-P₂, Ala, Phe and ATP in the three cellular populations allows us to conclude that the expression of pyruvate kinase is associated with the differentiation of these cells.Abbreviations GM-CFC granulocyte-macrophage colony forming cells - PK pyruvate kinase - CFU-E Colony Forming Units Erythroid - E_w Error weight - PEP phosphoenolpyruvate - Fru-1,6-P₂ fructose 1,6-bisphosphate - Ala L-alanine - Phe L-phenylanine - 5 GM granulocytemacrophage colonies obtained after 5 days incubation - 7 GM granulocyte-macrophage colonies obtained after 7 days incubation - h Hill coefficient - S_0,5 substrate concentration that yields half-maximal velocity     

Effects of glycerol on the in vitro stability and regulatory activation/inactivation of pyruvate,orthophosphate dikinase of Zea mays L.

Glycerol stabilizes the activity of pyruvate, orthophosphate dikinase extracted from darkened or illuminated maize leaves. It serves as a better protectant of activity than dithiothreitol for the active day-form and the glycerol concentration needed for full protection is inversely related to the level of protein. The night-form of the enzyme is also protected by glycerol not only against inactivation, but also against partial reactivation in storage. Glycerol does not prevent the P_i-dependent activation nor the ADP-dependent inactivation of pyruvate, orthophosphate dikinase, but the rates of both processes are substantially decreased. The ability of the inactive night-form for P_i-dependent activation is also sustained by glycerol for at least 2 h at 20°C, apparently through stabilization of the labile regulatory protein.Abbreviations BSA bovine serum albumin - G-6-P glucose-6-phosphate - MDH malate dehydrogenase - PCMB p-chloromercuribenzoate - PEP phosphoenolpyruvate - PEPCase phosphoenol-pyruvate carboxylase - PPDK pyruvate, orthophosphate dikinase - PVP polyvinylpyrrolidone     

Patterns of phosphoenolpyruvate carboxylase activity and cytosolic pH during light activation and dark deactivation in C3 and C4 plants

The rate and extent of light activation of PEPC may be used as another criterion to distinguish C₃ and C₄ plants. Light stimulated phosphoenolypyruvate carboxylase (PEPC) in leaf discs of C₄ plants, the activity being three times greater than that in the dark but stimulation of PEPC was limited about 30% over the dark-control in C₃ species. The light activation of PEPC in leaves of C₃ plants was complete within 10 min, while maximum activation in C₄ plants required illumination for more than 20 min, indicating that the relative pace of PEPC activation was slower in C₄ plants than in C₃ plants. Similarly, the dark-deactivation of the enzyme was also slower in leaves of C₄ than in C₃ species. The extent of PEPC stimulation in the alkaline pH range indicated that the dark-adapted form of the C₄ enzyme is very sensitive to changes in pH. The pH of cytosol-enriched cell sap extracted from illuminated leaves of C₄ plants was more alkaline than that of dark-adapted leaves. The extent of such light-dependent alkalization of cell sap was three times higher in C₄ leaves than in C₃ plants. The course of light-induced alkalization and dark-acidification of cytosol-enriched cell sap was markedly similar to the pattern of light activation and dark-deactivation of PEPC in Alternanthera pungens, a C₄ plant. Our report provides preliminary evidence that the photoactivation of PEPC in C₄ plants may be mediated at least partially by the modulation of cytosolic pH.Abbreviations CAM Crassulacean acid metabolism - G-6-P glucose-6-phosphate - PMSF phenylmethylsulfonyl fluoride - PEPC phosphoenolpyruvate carboxylase - PEPC-PK phosphoenolpyruvate ca carboxylase-protein kinase     

Assaying for pyruvate,orthophosphate dikinase activity: Necessary precautions with phosphoenolpyruvate carboxylase as coupling enzyme

Phosphoenolpyruvate carboxylase (EC 4.1.1.31), used as a coupling enzyme in the assay of the pyruvate, orthophosphate dikinase (EC 2.7.9.1) forward reaction, is a serious limiting factor for the overall rate when added at a level of 0.2–0.3 unit/ml of assay medium. Nonlimiting assay conditions are obtained by either increasing the level of the coupling enzyme to 3 units/ml or adding 6mM glucose-6-phosphate as an activator/stabilizer of phosphoenolpyruvate carboxylase.Abbreviations G-6-P glucose-6-phosphate - LDH lactate dehydrogenase - MDH malate dehydrogenase - PEP phosphoenolpyruvate - PEPCase phosphoenolpyruvate carboxylase - PVP polyvinylpyrrolidone - PPDK pyruvate, orthophosphate dikinase - U unit of enzyme activity (mol/min)     

Regulation of glycolysis and level of the Crassulacean acid metabolism

Glycolysis shows different patterns of operation and different control steps, depending on whether the level of Crassulacean acid metabolism (CAM) is low or high in the leaves of Kalanchoe blossfeldiana v.Poelln., when subjected to appropriate photoperiodic treatments: at a low level of CAM operation all the enzymes of glycolysis and phosphoenol pyruvate (PEP) carboxylase present a 12 h rhythm of capacity, resulting from the superposition of two 24h rhythms out of phase; phosphofructokinase appears to be the main regulation step; attainment of high CAM level involves (1) an increase in the peak of capacity occurring during the night of all the glycolytic enzymes, thus achieving an over-all 24h rhythm, in strict allometric coherence with the increase in PEP carboxylase capacity, (2) the establishment of different phase relationships between the rhythms of enzyme capacity, and (3) the control of three enzymic steps (phosphofructokinase, the group 3-P-glyceraldehyde dehydrogenase — 3-P-glycerate kinase, and PEP carboxylase). Results show that the hypothesis of allosteric regulation of phosphofructokinase (by PEP) and PEP carboxylase (by malate and glucose-6-P) cannot provide a complete explanation for the temporal organization of glycolysis and that changes in the phase relationships between the rhythms of enzyme capacity along the pathway and a strict correlation between the level of PEP carboxylase capacity and the levels of capacity of the glycolytic enzymes are important components of the regulation of glycolysis in relation to CAM.Abbreviations CAM crassulacean acid metabolism - F-6-P fructose-6-phosphate - F-bi-P fructose-1,6 biphosphate - G-3-PDH 3-phosphoglyceraldehyde dehydrogenase (NAD), EC 1.2.1.12 - G-6-P glucose-6-phosphate - GSH reduced glutathion - GDH glycerolphosphate dehydrogenase, EC 1.1.1.8 - PEP phosphoenol pyruvate - PEPC PEP carboxylase, EC 4.1.1.31 - PFK phosphofructokinase, EC 2.7.1.11 - 2-PGA 2-phosphoglycerate - 3-PGA 3-phosphoglycerate - PGM phosphoglycerate phosphomutase, EC 5.4.2.1 - T.P. triose phosphates - TPI triose phosphate isomerase, EC 5.3.1.1     

Altitude-related changes in activities of carbon metabolism enzymes in <Emphasis Type="Italic">Rumex nepalensis</Emphasis>

Activities of some enzymes related to carbon metabolism were studied in different ecotypes of Rumex nepalensis growing at 1 300, 2 250, and 3 250 m above mean sea level. Activities of ribulose-1,5-bisphosphate carboxylase/oxygenase, phosphoenolpyruvate carboxylase, aspartate aminotransferase, and glutamine synthetase increased with altitude, whereas activities of malate dehydrogenase, NAD-malic enzyme, and citrate synthase did not show a significant difference with change in altitude.     

Differential inhibition of photosynthesis under drought stress in <Emphasis Type="Italic">Flaveria</Emphasis> species with different degrees of development of the C<Subscript>4</Subscript> syndrome

The effect of drought stress (DS) on photosynthesis and photosynthesis-related enzyme activities was investigated in F. pringlei (C₃), F. floridana (C₃–C₄), F. brownii (C₄-like), and F. trinervia (C₄) species. Stomatal closure was observed in all species, probably being the main cause for the decline in photosynthesis in the C₃ species under ambient conditions. In vitro ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO) and stromal fructose 1,6-bisphosphatase (sFBP) activities were sufficient to interpret the net photosynthetic rates (P _N), but, from the decreases in P _N values under high CO₂ (C _a = 700 μmol mol^{− 1}) it is concluded that a decrease in the in vivo rate of the RuBPCO reaction may be an additional limiting factor under DS in the C₃ species. The observed decline in the photosynthesis capacity of the C₃–C₄ species is suggested to be associated both to in vivo decreases of RuBPCO activity and of the RuBP regeneration rate. The decline of the maximum P _N observed in the C₄-like species under DS was probably attributed to a decrease in maximum RuBPCO activity and/or to decrease of enzyme substrate (RuBP or PEP) regeneration rates. In the C₄ species, the decline of both in vivo photosynthesis and photosynthetic capacity could be due to in vivo inhibition of the phosphoenolpyruvate carboxylase (PEPC) by a twofold increase of the malate concentration observed in mesophyll cell extracts from DS plants.     

Urea and salt effects on enzymes from estivating and non-estivating amphibians

The effects of urea, cations (K⁺, NH₄, Na⁺, Cs⁺, Li⁺), and trimethylamines on the maximal activities and kinetic properties of pyruvate kinase (PK) and phosphofructokinase (PFK) from skeletal muscle, were analyzed in two anuran amphibians, an estivating species, the spadefoot toadScaphiopus couchii, and a semi-aquatic species, the leopard frogRana pipiens. Urea, which accumulates naturally to levels of 200–300 mM during estivation in toads, had only minor effects on the V_max, kinetic constants and pH curves of PK from either species and no effects on PFK V_max or kinetic constants. Trimethylamine oxide neither affected enzyme activity directly or changed enzyme response to urea. By contrast, high KCl (200 mM) lowered the V_max of toad PFK and of PK from both species and altered the K_m values for both substrates of frog PFK. Other cations were even more inhibitory; for example, the V_max of PK from either species was reduced by more than 80% by the addition of 200 mM NH₄Cl, NaCl, CsCi, or LiCl. High KCl also significantly changed the K_m values for substrates of toad lactate dehydrogenase and strongly reduced the V_max of glutamate dehydrogenase and NAD-dependent isocitrate dehydrogenase in both species whereas 300 mM urea had relatively little effect on these enzymes. The perturbing effect of urea on enzymes and the counteracting effect of trimethylamines that has been reported for elasmobranch fishes (that maintain high concentrations of both solutes naturally) does not appear to apply to amphibian enzymes. Rather, we found that urea is largely a non-perturbing solute for anuran enzymes (I₅₀ values were>1 M for both PK and PFK in both species) and we propose that its accumulation in high concentrations during estivation helps to minimize the increase in cellular ionic strength that would otherwise occur during desiccation and to alleviate the accompanying negative effects of high salt on individual enzyme activities and overall metabolic regulation.Abbreviations PFK 6-phosphofructo-1-kinase - PK pyruvate kinase     

Kinetic interactions of glycine with substrates and effectors of phosphenolpyruvate carboxylase from maize leaves

Glycine enhanced the sensitivity of maize phosphenolpyruvate carboxylase to the activator glucose 6-phosphate and reduced the sensitivity of the enzyme to the inhibitors malate and aspartate. The effects of glycine on the kinetic constants for these other effectors were greater than its effect on the K_m for substrate, raising the K_i(malate) 11-fold and reducing K_{a(glucose6-P)} 7-fold, while reducing the K_m(PEP) by 3-fold. Kinetically saturating levels of glycine and glucose 6-phosphate acted synergistically to raise K_i(malate) higher than that observed with either activator alone. Glycine and glucose 6-phosphate also synergistically reduced aspartate inhibition. Dual inhibitor analysis indicated that aspartate and malate bind in a mutually exclusive manner, and thus probably compete for the same inhibitor site. In contrast, the synergism between glycine and glucose 6-phosphate indicate that these activators bind at separate sites. Glycine also reduced the K_m(Mg) by 3-fold but had no significant effect on the K_m of bicarbonate.Abbreviation PEP phosphoenolpyruvate     

Effects of m-Cl-peroxy benzoic acid on glycolysis in Saccharomyces cerevisiae

Concentrations of m-Cl-peroxy benzoic acid (CPBA) higher than 0.1 mM decrease the ATP-content of Saccharomyces cerevisiae in the presence of glucose in 1 min to less than 10% of the initial value. In the absence of glucose, 1.0 mM CPBA is necessary for a similar effect. After the rapid loss of ATP in the first min in the presence of glucose caused by 0.2 mM CPBA, the ATP-content recovers to nearly the initial value after 10 min. Aerobic glucose consumption and ethanol formation from glucose are both completely inhibited by 1.0 mM CPBA. Assays of the activities of nine different enzymes of the glycolytic pathway as well as analysis of steady state concentrations of metabolites suggest that glyceraldehyde-3-phosphate dehydrogenase is the most sensitive enzyme of glucose fermentation. Phosphofructokinase and alcohol dehydrogenase are slightly less sensitive. Incubation for 1 or 10 min with concentrations of 0.05 to 0.5 mM CPBA causes a) inhibition of glyceraldehyde-3-phosphate dehydrogenase, b) decrease of the ATP-content and c) a decrease of the colony forming capacity. From these findings it is concluded that the disturbance of the ATP-producing glycolytic metabolism by inactivation of glyceraldehyde-3-phosphate dehydrogenase may be an explanation for cell death caused by CPBA.Abbreviations CPBA m-Chloro-peroxy benzoic acid - G-6-P glucose-6-phosphate - F-6-P fructose-6-phosphate - F-1,6-P₂ frnctose-1,6-bisphosphate - DAP dihydroxyacetone phosphate - GAP glyceraldehyde-3-phosphate - 2PGA 2-phosphoglycerate - PEP phosphoenol pyruvate - Pyr pyruvate - EtOH ethanol - PFK phosphofructokinase - GAPDH glyceraldehyde-3-phosphate dehydrogenase - ADH alcohol dehydrogenase Dedicated to Prof. Dr. Wolfgang Gerok at the occasion of his 60th birthday     

Evaluation of phosphorus starvation inducible genes relating to efficient phosphorus utilization in rice

Plants develop strategies to recycle phosphorus so that all organs receive adequate amounts of phosphorus, especially new growing organs. To evaluate the metabolic adaptation of rice plants under phosphorus deficient conditions, we selected several genes related to phosphorus utilization efficiency in the cell. Phosphoenolpyruvate carboxylase, triose phosphate translocator, phosphoenolpyruvate/phosphate translocator (PPT), pyruvate kinase, NAD dependent glyceraldehyde-3-phosphate dehydrogenase, and NADP dependent glyceraldehyde-3-phosphate dehydrogenase were selected because of their important roles in phosphorus utilization by the cell, and because they are part of the proposed bypass pathways by which the cells save phosphate. The most dramatic change was observed in the expression level of PPT (which transports phosphoenolpyruvate (PEP) from the cytosol into the chloroplast); thus we believe that PEP may play an important role in maintaining carbon metabolism under phosphate deficient conditions.     

A comparative study on the responses of the gills of two mudskippers to hypoxia and anoxia

A comparison of branchial enzyme profiles indicates that the gills of Periophthalmodon schlosseri would have a greater capacity for energy metabolism through glycolysis than those of Boleophthalmus boddaerti. Indeed, after exposure to hypoxia, or anoxia, there were significant increases in the lactate content in the gills of P. schlosseri. In addition, exposure to hypoxia or anoxia significantly lowered the glycogen level in the gills of this mudskipper. It can be deduced from these results that the glycolytic flux was increased to compensate for the decrease in ATP production through anaerobic glycolysis. Different from P. schlosseri, although there was an increase in lactate production in the gills of B. boddaerti exposed to hypoxia, there was no significant change in the branchial glycogen content, indicating that a reversed Pasteur effect might have occurred under such conditions. In contrast, anoxia induced an accumulation of lactate and a decrease in glycogen in the gills of B. boddaerti. Although lactate production in the gills of these mudskippers during hypoxia was inhibited by iodoacetate, the decreases in branchial glycogen contents could not account for the amounts of lactate formed. The branchial fructose-2,6-bisphosphate contents of these mudskippers exposed to hypoxia or anoxia decreased significantly, leaving phosphofructokinase and glycolytic rate responsive to cellular energy requirements under such conditions. The differences in response in the gills of B. boddaerti and P. schlosseri to hypoxia were possibly related to the distribution of phosphofructokinase between the free and bound states.Abbreviations ADP adenosine diphosphate - ALD aldolase - ALT alanine transaminase - AST aspartate transaminase - ATP adenosine triphosphate - CS citrate synthase - EDTA ethylenediaminetetra-acetic acid - EGTA ethylene glycol tetra-acetic acid - F6P fructose-6-phosphate - F-1,6-P₂ fructose-1,6-bisphosphate - F-2,6-P₂ fructose-2,6-bisphosphate - FBPase fructose-1,6-bisphosphatese - GAPDH glyceraldehyde-3-phosphate dehydrogenase - GDH glutamate dehydrogenase - -GDH -glycerophosphate dehydrogenase - GPase glycogen phosphorylase - HK hexokinase - HOAD 3-hydroxyacyl-CoA dehydrogenase - IDH isocitrate dehydrogenase - IOA iodoacetic acid - LDH lactate dehydrogenase - LO lactate oxidizing activity - MDH malate dehydrogenase - 3-PG 3-phosphoglyceric acid - PEP phosphoenolpyruvate - PEPCK phosphoenolpyruvate carboxykinase - PGI phosphoglucose isomerase - PGK phosphoglycerate kinase - PFK 6-phosphofructo-1-kinase - PIPES piperazine-N, N-bis-(2-ethanesulphonic acid) - PK pyruvate kinase - PMSF phenylmethylsulphonyl fluoride - PR pyrurate reducing activity - SE standard error - SW seawater - TPI triosephosphate isomerase