Effect of phosphoglycerate mutase deficiency on heterotrophic and autotrophic carbon metabolism of Alcaligenes eutrophus.

Mutants of Alcaligenes eutrophus were isolated on the basis of their inability to grow on succinate as the sole source of carbon and energy. The mutants also failed to grow on other gluconeogenic substrates, including pyruvate, acetate, and citrate. Simultaneously, they had lost their capability for autotrophic growth. The mutants grew, but slower than the wild type, on fructose or gluconate. Growth retardation on gluconate was more pronounced. The mutants lacked phosphoglycerate mutase activity, and spontaneous revertants of normal growth phenotype had regained the activity. The physiological characteristics of the mutants indicate the role of phosphoglycerate mutase in heterotrophic and autotrophic carbon metabolism of A. eutrophus. Although the enzyme is necessary for gluconeogenesis during heterotrophic growth on three- or four-carbon substrates, its glycolytic function is not essential for the catabolism of fructose or gluconate via the Entner-Doudoroff pathway. The enzyme is required during autotrophic growth as a catalyst in the biosynthetic route leading from glycerate 3-phosphate to pyruvate. It is suggested that the mutants accomplish the complete degradation of fructose and gluconate mutase lesion. The catabolically produced triose phosphates are converted to fructose 6-phosphate which is rechanneled into the Entner-Doudoroff pathway. This carbon recycling mechanism operates less effectively in mutant cells growing on gluconate.     

Alginic acid synthesis in Pseudomonas aeruginosa mutants defective in carbohydrate metabolism.

Mutant cells of mucoid Pseudomonas aeruginosa isolated from cystic fibrosis patients were examined for their ability to synthesize alginic acid in resting cell suspensions. Unlike the wild-type strain which synthesizes alginic acid from glycerol, fructose, mannitol, glucose, gluconate, glutamate, or succinate, mutants lacking specific enzymes of carbohydrate metabolism are uniquely impaired. A phosphoglucose isomerase mutant did not synthesize the polysaccharide from mannitol, nor did a glucose 6-phosphate dehydrogenase mutant synthesize the polysaccharide from mannitol or glucose. Mutants lacking the Entner-Doudoroff pathway dehydrase or aldolase failed to produce alginate from mannitol, glucose, or gluconate, as a 3-phosphoglycerate kinase or glyceraldehyde 3-phosphate dehydrogenase mutant failed to produce from glutamate or succinate. These results demonstrate the primary role of the Entner-Doudoroff pathway enzymes in the synthesis of alginate from glucose, mannitol, or gluconate and the role of glyceraldehyde 3-phosphate dehydrogenase reaction for the synthesis from gluconeogenic precursors such as glutamate. The virtual absence of any activity of phosphomannose isomerase in cell extracts of several independent mucoid bacteria and the impairment of alginate synthesis from mannitol in mutants lacking phosphoglucose isomerase or glucose 6-phosphate dehydrogenase rule out free mannose 6-phosphate as an intermediate in alginate biosynthesis.     

The route of ethanol formation in Zymomonas mobilis

1. Enzymic evidence supporting the operation of the Entner-Doudoroff pathway in the anaerobic conversion of glucose into ethanol and carbon dioxide by Zymomonas mobilis is presented. 2. Cell extracts catalysed the formation of equimolar amounts of pyruvate and glyceraldehyde 3-phosphate from 6-phosphogluconate. Evidence that 3-deoxy-2-oxo-6-phosphogluconate is an intermediate in this conversion was obtained. 3. Cell extracts of the organism contained the following enzymes: glucose 6-phosphate dehydrogenase (active with NAD and NADP), ethanol dehydrogenase (active with NAD), glyceraldehyde 3-phosphate dehydrogenase (active with NAD), hexokinase, gluconokinase, glucose dehydrogenase and pyruvate decarboxylase. Extracts also catalysed the overall conversion of glycerate 3-phosphate into pyruvate in the presence of ADP. 4. Gluconate dehydrogenase, fructose 1,6-diphosphate aldolase and NAD-NADP transhydrogenase were not detected. 5. It is suggested that NAD is the physiological electron carrier in the balanced oxidation-reduction involved in ethanol formation.     

Adenosine triphosphate-linked control of Pseudomonas aeruginosa glucose-6-phosphate dehydrogenase

Extracts of Pseudomonas aeruginosa (ATCC 7700) cells grown on glucose, gluconate, or glycerol had enzyme activities related to the Entner-Doudoroff pathway. These activities were present in no more than trace amounts when the bacteria were grown on succinate. Fructose-1,6-diphosphate aldolase could not be detected in extracts of the bacteria grown on any of the above carbon sources. Therefore, it appears that P. aeruginosa degrades glucose via an inducible Entner-Doudoroff pathway. The apparent absence of fructose-1,6-diphosphate aldolase in cells growing on succinate suggests that the bacteria can form hexose and pentose phosphates from succinate by an alternate route. d-Glucose-6-phosphate dehydrogenase, a branch-point enzyme of the Entner-Doudoroff pathway, was purified 50-fold from glucose-grown cells. Its molecular weight, estimated by sucrose density gradient centrifugation, was found to be approximately 190,000. The enzyme was strongly inhibited by adenosine triphosphate, guanosine triphosphate, and deoxyguanosine triphosphate, which decreased the apparent binding of glucose-6-phosphate to the enzyme. It is suggested that adenine nucleotide-linked control of glucose-6-phosphate dehydrogenase may regulate the overall catabolism of hexose phosphates and prevent their wasteful degradation under certain conditions requiring gluconeogenesis.     

Degradation of endogenous fructose during catabolism of sucrose and mannitol in halophilic archaebacteria

Metabolism of fructose arising endogenously from sucrose or mannitol was studied in halophilic archaebacteria Haloarcula vallismortis and Haloferax mediterranei. Activities of the enzymes of Embden-Meyerhof-Parnas (EMP) pathway, Entner-Doudoroff (ED) pathway and Pentose Phosphate (PP) pathway were examined in extracts of cells grown on sucrose or mannitol and compared to those grown on fructose and glucose. Sucrase and NAD-specific mannitol dehydrogenase were induced only when sucrose or mannitol respectively were the growth substrates. Endogenously arising fructose was metabolised in a manner similar to that for exogenously supplied fructose i.e. a modified EMP pathway initiated by ketohexokinase. While the enzymes for modified EMP pathway viz. ketohexokinase, 1-phosphofructokinase and fructose 1,6-bisphosphate aldolase were present under all growth conditions, their levels were elevated in presence of fructose. Besides, though fructose 1,6-bisphosphatase, phosphohexoseisomerase and glucose 6-phosphate dehydrogenase were present, the absence of 6-phosphogluconate dehydratase precluded routing of fructose through ED pathway, or through PP pathway directly as 6-phosphogluconate dehydrogenase was lacking. Fructose 1,6-bisphosphatase plays the unusual role of a catabolic enzyme in supporting the non-oxidative part of PP pathway. However the presence of constitutive levels of glucose dehydrogenase and 2-keto 3-deoxy 6-phosphogluconate aldolase when glucose or sucrose were growth substrates suggested that glucose breakdown took place via the modified ED pathway.Abbreviations EMP Embden Meyerhof Parnas - ED Entner Doudoroff - PP pentose phosphate - KHK ketohexokinase - 1-PFK 1-phosphofructokinase - PEP-PTS phosphoenolpyruvate phosphotransferase - 6-PFK 6-phosphofructokinase - FBPase fructose 1,6-bisphosphatase - PHI phosphohexoseisomerase - G6P-DH glucose 6-phosphate dehydrogenase - 6PG-DH 6-phosphogluconate dehydrogenase - GAPDH glyceraldehyde 3-phosphate dehydrogenase - FIP fructose 1-phosphate - GSH reduced glutathione - 2-ME -mercaptoethanol - FBP fructose 1,6-bisphosphate - KDPG 2-keto 3-deoxy 6-phosphogluconate - F6P fructose 6-phosphatez     

Glucose and Gluconate Metabolism in a Mutant of Escherichia coli Lacking Gluconate-6-phosphate Dehydrase

A mutant lacking gluconate-6-phosphate dehydrase (the first enzyme of the Entner-Doudoroff pathway) was isolated after ethyl methane sulfonate mutagenesis of Escherichia coli. Other enzymes of gluconate metabolism (gluconokinase, gluconate-6-phosphate dehydrogenase, and 2-keto-3-deoxygluconate-6-phosphate aldolase) were present in the mutant. When the mutant was grown on gluconate-1-(14)C, alanine isolated from protein was unlabeled, showing that the dehydrase was absent in vivo and that the sole pathway of gluconate metabolism in the mutant was the hexose monophosphate shunt. The mutant grew on gluconate with a doubling time of 155 min, compared with the parent strain's 56 min. On glucose and fructose it grew with normal doubling times. Thus, in E. coli, the Entner-Doudoroff pathway is used for gluconate metabolism but not for glucose metabolism.     

The enzymes of the classical pentose phosphate pathway display differential activities in procyclic and bloodstream forms of Trypanosoma brucei

The specific activities of each of the enzymes of the classical pentose phosphate pathway have been determined in both cultured procyclic and bloodstream forms of Trypanosoma brucei. Both forms contained glucose-6-phosphate dehydrogenase (EC 1.1.1.49), 6-phosphogluconolactonase (EC 3.1.1.31), 6-phosphogluconate dehydrogenase (EC 1.1.1.44), ribose-5-phosphate isomerase (EC 5.3.1.6) and transaldolase (EC 2.2.1.2). However, ribulose-5-phosphate 3'-epimerase (EC 5.1.3.1) and transketolase (EC 2.2.1.1) activities were detectable only in procyclic forms. These results clearly demonstrate that both forms of T. brucei can metabolize glucose via the oxidative segment of the classical pentose phosphate pathway in order to produce D-ribose-5-phosphate for the synthesis of nucleic acids and reduced NADP for other synthetic reactions. However, only procyclic forms are capable of using the non-oxidative segment of the classical pentose phosphate pathway to cycle carbon between pentose and hexose phosphates in order to produce D-glyceraldehyde 3-phosphate as a net product of the pathway. Both forms lack the key gluconeogenic enzyme, fructose-bisphosphatase (EC 3.1.3.11). Consequently, neither form should be able to engage in gluconeogenesis nor should procyclic forms be able to return any of the glyceraldehyde 3-phosphate produced in the pentose phosphate pathway to glucose 6-phosphate. This last specific metabolic arrangement and the restriction of all but the terminal steps of glycolysis to the glycosome may be the observations required to explain the presence of distinct cytosolic and glycosomal isoenzymes of glyceraldehyde-3-phosphate dehydrogenase and phosphoglycerate kinase. These same observations also may provide the basis for explaining the presence of cytosolic hexokinase and phosphoglucose isomerase without the presence of any cytosolic phosphofructokinase activity. The key enzymes of the Entner-Doudoroff pathway, 6-phosphogluconate dehydratase (EC 4.2.1.12) and 2-keto-3-deoxy-6-phosphogluconate aldolase (EC 4.1.2.14) were not detected in either procyclic or bloodstream forms of T. brucei.     

Carbon Metabolism Enzymes of Rhizobium tropici Cultures and Bacteroids

We determined the activities of selected enzymes involved in carbon metabolism in free-living cells of Rhizobium tropici CFN299 grown in minimal medium with different carbon sources and in bacteroids of the same strain. The set of enzymatic activities in sucrose-grown cells suggests that the pentose phosphate pathway, with the participation of the Entner-Doudoroff pathway, is probably the primary route for sugar catabolism. In glutamate- and malate-grown cells, high activities of the gluconeogenic enzymes (phosphoenolpyruvate carboxykinase, fructose-6-phosphate aldolase, and fructose bisphosphatase) were detected. In bacteroids, isolated in Percoll gradients, the levels of activity for many of the enzymes measured were similar to those of malate-grown cells, except that higher activities of glucokinase, glucose-6-phosphate dehydrogenase, and NAD-dependent phosphogluconate dehydrogenase were detected. Phosphoglucomutase and UDP glucose pyrophosphorylase showed high and constant levels under all growth conditions and in bacteroids.     

Utilization of Glucose by Clostridium thermocellum: Presence of Glucokinase and Other Glycolytic Enzymes in Cell Extracts

Clostridium thermocellum was shown to ferment glucose in a medium containing salts and 0.5% yeast extract. An active glucokinase was obtained with improved conditions for growth, assay, and preparation of cell extracts. Cell extracts appear to contain a glucokinase inhibitor that interferes with the assays at high protein concentrations. Glucokinase activity is stimulated about 60% by pretreatment with dithiothreitol. Little or no fructokinase or mannokinase activity was detected in cell extracts. The absence of glucokinase in mannitol-grown cells, the increase in glucokinase activity upon incubation of cell suspensions with glucose, and the lack of increase in activity when chloramphenical is added are evidence that glucokinase is an inducible enzyme. The following enzymes were detected in cell extracts (the enzyme activities are shown in parentheses are micromoles per minute per milligram or protein at 27 C): glucokinase (0.48), phosphoglucose isomerase (0.73), fructose 6-phosphate kinase (0.24), fructose diphosphate aldolase (0.59), glyceraldehyde 3-phosphate dehydrogenase (0.53), triose phosphate isomerase (0.13), phosphoglycerate kinase (0.20), phosphoglycerate mutase (0.20), enolase (0.28), pyruvic kinase (0.13), and lactic dehydrogenase (0.13). Glucose 6-phosphate dehydrogenase activity was absent or very low (0.0002) and 6-phosphogluconate dehydrogenase activity also was relatively low (0.015). From these data, it is proposed that carbohydrate metabolism in C. thermocellum proceeds by the Embden-Meyerhof pathway.     

Mode of action of alpha-chlorohydrin as a male anti-fertility agent. Inhibition of the metabolism of ram spermatozoa by alpha-chlorohydrin and location of block in glycolysis

1. The effect of alpha-chlorohydrin on the metabolism of glycolytic and tricarboxylate-cycle substrates by ram spermatozoa was investigated. The utilization and oxidation of fructose and triose phosphate were much more sensitive to inhibition by alpha-chlorohydrin (0.1-1.0mm) than lactate or pyruvate. Inhibition of glycolysis by alpha-chlorohydrin is concluded to be between triose phosphate and pyruvate formation. Oxidation of glycerol was not as severely inhibited as that of the triose phosphate. This unexpected finding can be explained in terms of competition between glycerol and alpha-chlorohydrin. A second, much less sensitive site, of alpha-chlorohydrin inhibition appears to be associated with production of acetyl-CoA from exogenous and endogenous fatty acids. 2. Measurement of the glycolytic intermediates after incubation of spermatozoal suspensions with 15mm-fructose in the presence of 3mm-alpha-chlorohydrin showed a ;block' in the conversion of glyceraldehyde 3-phosphate into 3-phosphoglycerate. alpha-Chlorohydrin also caused conversion of most of the ATP in spermatozoa into AMP. After incubation with 3mm-alpha-chlorohydrin, glyceraldehyde 3-phosphate dehydrogenase and triose phosphate isomerase activities were decreased by approx. 90% and 80% respectively, and in some experiments aldolase was also inhibited. Other glycolytic enzymes were not affected by a low concentration (0.3mm) of alpha-chlorohydrin. Loss of motility of spermatozoa paralleled the decrease in glyceraldehyde 3-phosphate dehydrogenase activity. alpha-Chlorohydrin, however, did not inhibit glyceraldehyde 3-phosphate dehydrogenase or triose phosphate isomerase in sonicated enzyme preparations when added to the assay cuvette. 3. Measurement of intermediates and glycolytic enzymes in ejaculated spermatozoa before, during and after injection of rams with alpha-chlorohydrin (25mg/kg body wt.) confirmed a severe block in glycolysis in vivo at the site of triose phosphate conversion into 3-phosphoglycerate within 24h of the first injection. Glyceraldehyde 3-phosphate dehydrogenase activity was no longer detectable and both aldolase and triose phosphate isomerase were severely inhibited. Spermatozoal ATP decreased by 92% at this time, being quantitatively converted into AMP. At 1 month after injection of alpha-chlorohydrin glycolytic intermediate concentrations returned to normal in the spermatozoa but ATP was still only 38% of the pre-injection concentration. Motility of spermatozoa was, however, as good as during the pre-injection period. The activity of the inhibited enzymes also returned to normal during the recovery period and 26 days after injection were close to pre-injection values. 4. An unknown metabolic product of alpha-chlorohydrin is suggested to inhibit glyceraldehyde 3-phosphate dehydrogenase and triose phosphate isomerase of spermatozoa. This results in a lower ATP content, motility and fertility of the spermatozoa. Glycidol was shown not to be an active intermediate of alpha-chlorohydrin in vitro.     

The pentose phosphate pathway of glucose metabolism. Measurement of the non-oxidative reactions of the cycle

Methods for the quantitative determination of ribose 5-phosphate isomerase, ribulose 5-phosphate 3-epimerase, transketolase and transaldolase in tissue extracts are described. The determinations depend on the measurement of glyceraldehyde 3-phosphate by using the coupled system triose phosphate isomerase, α-glycero-phosphate dehydrogenase and NADH. By using additional purified enzymes transketolase, ribose 5-phosphate isomerase and ribulose 5-phosphate epimerase conditions could be arranged so that each enzyme in turn was made rate-limiting in the overall system. Transaldolase was measured with fructose 6-phosphate and erythrose 4-phosphate as substrates, and again glyceraldehyde 3-phosphate was measured by using the same coupled system. Measurements of the activities of the non-oxidative reactions of the pentose phosphate pathway were made in a variety of tissues and the values compared with those of the two oxidative steps catalysed by glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase.     

Enzymes involved in the formation of glycerol 3-phosphate and the by-products dihydroxyacetone and glycerol in Zymomonas mobilis

Abstract In Zymomonas mobilis a novel pathway for the formation of glycerol 3-phosphate was identified by enzymatic studies and nuclear magnetic resonance spectroscopy. This pathway branches off from the Entner-Doudoroff pathway at the intermediate glyceraldehyde 3-phosphate and proceedes via dihydroxyacetone phosphate, dihydroxyacetone, glycerol to glycerol 3-phosphate. The reaction sequence is catalyzed by the enzymes triosephosphate isomerase (0.4 U (mg protein)⁻¹), dihydroxyacetone phosphatase (0.31 U (mg protein)⁻¹), dihydroxyacetone reductase (0.25 U (mg protein)⁻¹), and glycerokinase (0.08 mU (mg protein)⁻¹), respectively. The action of a postulated aldolase catalyzing the cleavage of fructose 6-phosphate to dihydroxyacetone and glyceraldehyde 3-phosphate could be excluded.     

Heterotrophic Metabolism of the Chemolithotroph Thiobacillus ferrooxidans

Glucose-6-phosphate dehydrogenase and the enzymes of the Entner-Doudoroff pathway, 6-phosphogluconate dehydrase and 2-keto-3-deoxy-6-phosphogluconate aldolase (assayed together), are induced during heterotrophic growth of Thiobacillus ferrooxidans on an iron-glucose-supplemented medium or on glucose alone. By contrast, autotrophic cells (iron-grown) contain low levels of these enzymes. Fructose 1, 6-diphosphate aldolase, an enzyme of the Embden-Meyerhof pathway, is present at low levels irrespective of the growth medium, suggesting that this enzyme is not involved in energy-yielding reactions but merely provides intermediates for biosynthesis. The Entner-Doudoroff and pentose-phosphate pathways are the principle means through which glucose is dissimilated and is presumed to be concerned with energy production. Isotopic studies showed that a high rate of CO(2) formation from specifically labeled glucose came from carbon atoms 1 and 4. An unexpectedly high rate of evolution of CO(2) also came from carbon 6, suggesting that the triose phosphate formed during glucose breakdown and specifically as a result of 2-keto-3-deoxy-6-phosphogluconate aldolase activity, was metabolized via some unorthodox metabolic route. Cells grown in the iron-supplemented and glucose-salts media have a complete tricarboxylic acid cycle, whereas autotrophically grown T. ferrooxidans lacked both alpha-ketoglutarate dehydrogenase and reduced nicotinamide adenine dinucleotide oxidase. Two isocitrate dehydrogenases [nicotinamide adenine dinucleotide (NAD) and NAD phosphate (NADP) specific] were present. NAD-linked enzyme was constitutive, whereas the NADP-linked enzyme was induced upon adaptation of autotrophic cells to heterotrophic growth.     

Glucolysis in Pseudomonas putida: Physiological Role of Alternative Routes from the Analysis of Defective Mutants

A number of mutants in which glucolysis is impaired have been isolated from Pseudomonas putida. The study of their behavior shows that this organism possesses a single glucolytic pathway with physiological significance. The first step of the pathway consists in the oxidation of glucose into gluconate. Two proteins with glucose dehydrogenase activity appear to exist in P. putida but the reasons for this duplicity are not clear. The process continues with the formation of 2-ketogluconate which is in turn converted into gluconate-6-phosphate. This is proved by the fact that mutants unable to form gluconate-6-phosphate from 2-ketogluconate show extremely slow growth on glucose or gluconate (generation times are increased more than 100 times). Other possible routes for the conversion of glucose into gluconate-6-phosphate, the glucose-6-phosphate pathway, or the direct phosphorylation of the gluconate formed by glucose oxidation are only minor shunts in P. putida. The Entner-Doudoroff enzymes, which catalyze the conversion of gluconate-6-phosphate into pyruvate and triosephosphate, appear to be essential to grow on glucose and also on gluconate and 2-ketogluconate. A significative role of the pentose route in the catabolism of these substrates is not apparent from this study. In contrast, P. putida strains showing no activity of the Entner-Doudoroff enzymes grow readily on fructose, although there is evidence that this hexose is at least partially catabolized via gluconate-6-phosphate.     

Enzymes involved in the utilization of carbohydrates by two strains of myxobacteria

Summary Extracts prepared from two strains of myxobacteria, grown with glucose as the primary carbon source, have been shown to contain a hexokinase that phosphorylates glucose to a greater extent than fructose. Phosphohexose isomerase, 6-phosphofructokinase, aldolase and a DPN-specific triose phosphate dehydrogenase are also present. In addition a TPN-specific glucose-6-phosphate dehydrogenase and a system catalyzing the reduction of DPN in the presence of 6-phosphogluconate have also been demonstrated.Supported in part by State of Washington funds for medical and biological research, and in part by the Office of Naval Research under Contract N80nr-520/III with the University of Washington.     

Regulation of alternate peripheral pathways of glucose catabolism during aerobic and anaerobic growth of Pseudomonas aeruginosa.

Glucose may be converted to 6-phosphogluconate by alternate pathways in Pseudomonas aeruginosa. Glucose is phosphorylated to glucose-6-phosphate, which is oxidized to 6-phosphogluconate during anaerobic growth when nitrate is used as respiratory electron acceptor. Mutant cells lacking glucose-6-phosphate dehydrogenase are unable to catabolize glucose under these conditions. The mutant cells utilize glucose as effectively as do wild-type cells in the presence of oxygen; under these conditions, glucose is utilized via direct oxidation to gluconate, which is converted to 6-phosphogluconate. The membrane-associated glucose dehydrogenase activity was not formed during anaerobic growth with glucose. Gluconate, the product of the enzyme, appeared to be the inducer of the gluconate transport system, gluconokinase, and membrane-associated gluconate dehydrogenase. 6-Phosphogluconate is probably the physiological inducer of glucokinase, glucose-6-phosphate dehydrogenase, and the dehydratase and aldolase of the Entner-Doudoroff pathway. Nitrate-linked respiration is required for the anaerobic uptake of glucose and gluconate by independently regulated transport systems in cells grown under denitrifying conditions.     

Inhibition of the glycolytic pathway by methylglyoxal in human platelets

The incubation of human platelets with methylglyoxal and glucose produces a rapid transformation of the ketoaldehyde to D-lactate by the glyoxalase system and a partial reduction in GSH. Glucose utilization is affected at the level of the glycolytic pathway. No effect of the ketoaldehyde on glycogenolysis and glucose oxidation through the hexose monophosphate shunt was demonstrated. Phosphofructokinase, fructose 1,6 diphosphate (F1, 6DP) aldolase, glyceraldehyde 3-phosphate dehydrogenase and 3-phosphoglycerate mutase were mostly inhibited by methylglyoxal. A decrease in lactate and pyruvate formation and an accumulation of some glycolytic intermediates (fructose 1,6 diphosphate, dihydroxyacetone phosphate, 3-phosphoglycerate) was observed. Moreover methylglyoxal induced a fall in the metabolic ATP concentration. Since methylglyoxal is an intermediate of the glycolytic bypass system from dihydroxyacetone phosphate to D-lactate, it may be assumed that ketoaldehyde exerts a regulating effect on triose metabolism.     

Chloroplast Aldolase is Controlled by a Nuclear Gene

Variant chloroplast fructose 1,6-diphosphate aldolases were found in Pisum sativum when 10 commercial varieties were examined for electrophoretically distinct species of chloroplast triose phosphate isomerase, phosphoglyceric acid kinase, glyceraldehyde 3-phosphate dehydrogenase, and aldolase. When reciprocal crosses are made, both aldolases appear in individuals in the F(1) generation. Backcrossing gives offspring having aldolases characteristic of the homozygous or of the heterozygous parent; the inheritance is therefore not maternal but Mendelian. Clearly this chloroplast reductive pentose phosphate cycle enzyme is under nuclear gene control in P. sativum.     

Sugar metabolism in the extremely halophilic bacterium Salinibacter ruber

Growth of Salinibacter ruber, a red, extremely halophilic bacterium phylogenetically affiliated with the Flavobacterium/Cytophaga branch of the domain Bacteria, is stimulated by a small number of sugars (glucose, maltose, starch at 1 g l(-1)). Glucose consumption starts after other substrates have been depleted. Glucose metabolism proceeds via a constitutive, salt-inhibited hexokinase and a constitutive salt-dependent nicotinamide adenine dinucleotide phosphate (NADP)-linked glucose-6-phosphate dehydrogenase. Glucose dehydrogenase and fructose-1,6-bisphosphate aldolase activity could not be detected. It is therefore suggested that Salinibacter metabolizes glucose by the classic Entner-Doudoroff pathway and not by the Embden-Meyerhof glycolytic pathway or by the modified Entner-Doudoroff pathway present in halophilic Archaea of the family Halobacteriaceae, in which the phosphorylation step is postponed. However, activity of 2-keto-3-deoxy-6-phosphogluconate aldolase could not be detected in extracts of Salinibacter cells, whether or not grown in the presence of glucose.     

Glucose fermentation to acetate and alanine in resting cell suspensions of Pyrococcus furiosus: Proposal of a novel glycolytic pathway based on 13C labelling data and enzyme activities

Abstract Suspensions of maltose-grown cells of the hyperthermophilic archaeon Pyrococcus furiosus , when incubated at 90°C with 35 mM [1-¹³C]glucose or [3-¹³C]glucose, consumed glucose at a rate of about 10 nmol min⁻¹ (mg protein)⁻¹. Acetate (10 mM), alanine (3 mM), CO₂ and H₂ were the fermentation products. The ¹³C-labelling pattern in alamine and acetate were analyzed. With [1-¹³C]glucose the methyl group of both alanine and acetate was labelled; with [3-¹³C]glucose only the carboxyl group of alanine was labelled whereas acetate was unlabelled. Extracts of maltose-grown cells contained glucose isomerase (12.8 U mg⁻¹, 100°C), ketohexokinase (0.23 U mg⁻¹, 100°C), and fructose 1-phosphate aldolase (0.06 U mg^{−1, 100°C). Enzymes catalyzing the formation of fructose 1,6-bisphosphate from fructose 1-phosphate or fructose 6-phosphate could not be detected. As publihed previously by our group and other authors P. furiosus also contains enzymes of glyceraldehyde conversion to 2-phosphoglycerate according to a non-phosphorylated Entner-Doudoroff pathway, of dihydroxyacetone phosphate conversion to 2-phosphoglycerate according to the Embden-Meyerhof pathway, and of 2-phosphoglycerate conversion - via pyruvate - to acetate and alanine. Based on the enzyme activities in P. furiosus , the following pathway for glucose degradation to alanine and acetate in cell suspensions is proposed which can explain the [13}C]glucose labelling data: glucose→ fructose → fructose 1- phosphate → dihydroxyacetone phosphate + glyceraldehyde and further conversion of both trioses to alanine and acetate via pyruvate.