Glucose Metabolism in Neisseria gonorrhoeae

The metabolism of glucose was examined in several clinical isolates of Neisseria gonorrhoeae. Radiorespirometric studies revealed that growing cells metabolized glucose by a combination on the Entner-Doudoroff and pentose phosphate pathways. A portion of the glyceraldehyde-3-phosphate formed via the Entner-Doudoroff pathway was recycled by conversion to glucose-6-phosphate. Subsequent catabolism of this glucose-6-phosphate by either the Entner-Doudoroff or pentose phosphate pathways yielded CO(2) from the original C6 of glucose. Enzyme analyses confirmed the presence of all enzymes of the Entner-Doudoroff, pentose phosphate, and Embden-Meyerhof-Parnas pathways. There was always a high specific activity of glucose-6-phosphate dehydrogenase (EC 1.1.1.49) relative to that of 6-phosphogluconate dehydrogenase (EC 1.1.1.44). The glucose-6-phosphate dehydrogenase utilized either nicotinamide adenine dinucleotide phosphate or nicotinamide adenine dinucleotide as electron acceptor. Acetate was the only detectable nongaseous end product of glucose metabolism. Following the disappearance of glucose, acetate was metabolized by the tricarboxylic acid cycle as evidenced by the preferential oxidation of [1-(14)C]acetate over that of [2-(14)C]acetate. When an aerobically grown log-phase culture was subjected to anaerobic conditions, lactate and acetate were formed from glucose. Radiorespirometric studies showed that under these conditions, glucose was dissimilated entirely by the Entner-Doudoroff pathway. Further studies determined that this anaerobic dissimilation of glucose was not growth dependent.     

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.     

Regulation of Glucose Metabolism in Thiobacillus intermedius

Glucose-yeast extract or glucose-casein hydrolysate-grown Thiobacillus intermedius cells, which use glucose for energy generation, possess high specific activities of the Entner-Doudoroff pathway and related enzymes, 6-phosphogluconate dehydrase, 2-keto-3-deoxy-6-phosphogluconate aldolase, glucokinase, and glucose-6-phosphate dehydrogenase, but low activities of enzymes unique to the pentose shunt and Embden-Meyerhof pathways. Although the synthesis of the latter enzymes remains largely unaffected by the growth environment, that of the former is stimulated by glucose. Radiorespirometric measurements demonstrate an early and parallel respiration of glucose carbon atoms one and four in glucose-casein hydrolysate broth. It is concluded that the Entner-Doudoroff pathway performs an energetic role in glucose metabolism by T. intermedius with the pentose shunt and Embden-Meyerhof pathways functioning mainly in biosynthesis. The presence of thiosulfate in the growth medium inhibits the synthesis of the Entner-Doudoroff pathway and related enzymes. In addition, both thiosulfate and glucose inhibit the synthesis of the Krebs cycle enzymes, nicotinamide adenine dinucleotide phosphate-linked isocitrate and alpha-ketoglutarate dehydrogenases. Thus, repression of enzymes is of significance in the adaptation of T. intermedius to its nutritional environment. The activity of glucose-6-phosphate dehydrogenase of T. intermedius is inhibited by adenosine triphosphate. Such a control could afford the organism a mechanism to regulate the flow of glucose into major energetic and biosynthetic routes.     

Some effects of mannitol on the glucose metabolism of two derivatives of a strain ofRhizobium japonicum differing in mannitol dehydrogenase

The effects of mannitol were investigated by comparing some metabolic features in colonial derivatives, I-110 and L1-110, ofRhizobium japonicum strain 3IIb110, grown either on glucose alone (G-cells) or in glucose media supplemented with mannitol (GM-cells). The polyol stimulated the synthesis of not only mannitol dehydrogenase, which is active in derivative L1-110, but also the nicotinamide adenine dinucleotide (NAD)-linked 6-phosphogluconate (6-PG) dehydrogenase (EC 1.1.1.43). As revealed by radiorespirometry, when GM-cells were allowed to metabolize glucose, they produced relatively more CO₂ from the first and sixth carbons of the sugar than G-cells did. This finding is evidence that NAD-linked 6-PG dehydrogenase might initiate an unknown pathway different from the hexose cycle and the pentose phosphate (PP) pathway. Mannitol exerted no allosteric control on the oxygen consumption and the glucose transport systems. Active uptake of the polyol was correlated with the presence of mannitol dehydrogenase (EC 1.1.1.67); it did not hinder the transport of glucose even though both systems derive their energy for active transport from a common source presumptively characterized as the energized membrane state. Mannitol, however, suppressed by two- or threefold the glucose uptake system. Addition of the polyol to the cell suspensions of both colonial types ofR. japonicum metabolizing glucose caused an immediate 40–50% drop of adenosine triphosphate (ATP) concentrations, owing in part to the mannitol kinase reaction. Type I-110 failed to overcome this reduction of ATP levels, and low growth rates could results. In contrast, type L1-110 offsets the reduction of ATP concentration by oxidizing mannitol as an additional source of energy through mannitol dehydrogenase, fructokinase, and a sequence of glycolytic reactions. The polyol also induced type L1-110 to produce extracellular slimy materials that, apparently, harbor amounts of ATP and proteins.     

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 Catabolism in Rhizobium japonicum

Glucose catabolism in Rhizobium japonicum ATCC 10324 was investigated by the radiorespirometric method and by assaying for key enzymes of the major energy-yielding pathways. Specifically labeled glucose gave the following results for resting cells, with values expressed as per cent (14)CO(2) evolution: C-1=59%, C-2=51%, C-3=45%, C-4=59%, and C-6=43%. These values indicate that glucose was degraded by the Entner-Doudoroff pathway alone. Cells which grew in glucose-yeast extract-salts medium gave essentially the same pattern except for retardation of the C-6 carbon. The rates were: C-1=54%, C-2=42%, C-3=51%, C-4=59%, and C-6=32%. Hexokinase, glucose-6-phosphate dehydrogenase, transketolase, and an enzyme system which produces pyruvate from 6-phosphogluconate were found to be present in these cells. No 6-phosphogluconate dehydrogenase was detected. Oxidation of specifically labeled pyruvate gave the following (14)CO(2) evolution pattern: C-1=78%, C-2=48%, and C-3=37%; the pattern from acetate was C-1=73%; and C-2=56%. Oxidation of glutamate showed the preferential rate of (14)CO(2) evolution to be C-1 > C-2=C-5 > C-3, 4, whereas a higher yield of (14)CO(2) was obtained from the C-1 and C-4 carbons of succinate than from the C-2 and C-3 carbons. These data are consistent with the operation of the Entner-Doudoroff pathway and tricarboxylic acid cycle as the catabolic pathways of glucose oxidation in R. japonicum.     

Glucose Catabolism in Strains of Acidophilic, Heterotrophic Bacteria

Pathways of glucose catabolism, potentially operational in six strains of obligately aerobic, acidophilic bacteria, including Acidiphilium cryptum strain Lhet2, were investigated by short-term radiorespirometry and enzyme assays. Short-term radiorespirometry was conducted at pH 3.0 with specifically labeled [¹⁴C]glucose. The high rate and yield of C-1 oxidized to CO₂ indicated that the Entner-Doudoroff, pentose phosphate, or both pathways were operational in all strains. Apparent nonequivalent yields of CO₂ from C-1 and estimated CO₂ from C-4 (C-1 > C-4) were suggestive of simultaneous glucose catabolism by both pathways in all strains tested. Variation in the relative contribution of the two pathways of glucose catabolism appears to account for observed strain differences. Calculation of the actual percent pathway participation was not feasible. Enzyme assays were completed with crude extracts of glucose-grown cells to substantiate the results obtained by radiorespirometry. The key enzymes of the pentose phosphate pathway (6-phosphogluconate dehydrogenase) and the Entner-Doudoroff pathway (2-keto-3-deoxy-6-phosphogluconate aldolase and 6-phosphogluconate dehydrase) were present in all strains examined (PW2, Lhet2, KLB, OP, and QBP). However, none of the strains exhibited detectable levels of the key enzyme of the Embden-Meyerhof-Parnas pathway, 6-phosphofructokinase. All strains contained glucose-6-phosphate dehydrogenase and fructose bisphosphate aldolase. The results of the enzyme study supported the contention that the pentose phosphate and Entner-Doudoroff pathways are operational for glucose catabolism in the acidophilic heterotrophs, and that the Embden-Meyerhof-Parnas pathway is apparently absent.     

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     

Enzymes of glucose metabolism in Frankia sp.

Enzymes of glucose metabolism were assayed in crude cell extracts of Frankia strains HFPArI3 and HFPCcI2 as well as in isolated vesicle clusters from Alnus rubra root nodules. Activities of the Embden-Meyerhof-Parnas pathway enzymes glucokinase, phosphofructokinase, and pyruvate kinase were found in Frankia strain HFPArI3 and glucokinase and pyruvate kinase were found in Frankia strain HFPCcI2 and in the vesicle clusters. An NADP+-linked glucose 6-phosphate dehydrogenase and an NAD-linked 6-phosphogluconate dehydrogenase were found in all of the extracts, although the role of these enzymes is unclear. No NADP+-linked 6-phosphogluconate dehydrogenase was found. Both dehydrogenases were inhibited by adenosine 5-triphosphate, and the apparent Km's for glucose 6-phosphate and 6-phosphogluconate were 6.86 X 10(-4) and 7.0 X 10(-5) M, respectively. In addition to the enzymes mentioned above, an NADP+-linked malic enzyme was detected in the pure cultures but not in the vesicle clusters. In contrast, however, the vesicle clusters had activity of an NAD-linked malic enzyme. The possibility that this enzyme resulted from contamination from plant mitochondria trapped in the vesicle clusters could not be discounted. None of the extracts showed activities of the Entner-Doudoroff enzymes or the gluconate metabolism enzymes gluconate dehydrogenase or gluconokinase. Propionate- versus trehalose-grown cultures of strain HFPArI3 showed similar activities of most enzymes except malic enzyme, which was higher in the cultures grown on the organic acid. Nitrogen-fixing cultures of strain HFPArI3 showed higher specific activities of glucose 6-phosphate and 6-phosphogluconate dehydrogenases and phosphofructokinase than ammonia-grown cultures.     

Schistosoma mansoni and Schistosoma japonicum: comparison of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase activities in adults.

Glucose-6-phosphate dehydrogenase activity in paired adult Schistosoma mansoni is about twice as great as in paired adult Schistosoma japonicum. 2. 6-phosphogluconate dehydrogenase activity accounts for 25.8% of the measured production of reduced nicotinamide adenine dinucleotide phosphate (NADPH) in S. japonicum but only 8.6% of the measured production of NADPH in S. mansoni. 3. These data suggest a species difference in 6-phosphogluconate metabolism.     

Fructose catabolism in Azospirillum brasilense and Azospirillum lipoferum.

The pathways for catabolism of fructose were investigated in the type strains of Azospirillum lipoferum and Azospirillum brasilense grown aerobically with (NH4)2SO4 as the nitrogen source. When grown on fructose, the former species possessed a complete Entner-Doudoroff pathway, whereas the latter species lacked activity for glucose-6-phosphate dehydrogenase. Both species possessed a complete catabolic Embden-Meyerhof-Parnas pathway. Neither species possessed the key enzyme of the hexose monophosphate pathway, 6-phosphogluconate dehydrogenase. Both species could phosphorylate fructose to fructose-1-phosphate by means of a phosphoenolpyruvate-phosphotransferase system, and high activities of 1-phosphofructokinase occurred. Both species possessed glucokinase activity, but only A. lipoferum had hexokinase activity; moreover, the cells of A. brasilense were nearly impermeable to glucose, accounting for the inability of this species to grow on glucose. Both species possessed pyruvate dehydrogenase, a complete tricarboxylic acid cycle, a glyoxylate shunt, and malic enzyme. Analysis of the acidic end products for both species indicated the formation of only small amounts of various organic acids, and most of the titratable acidity was due to utilization of the ammonium ions of the medium. Gluconic acid was not formed during growth of either species on fructose but was detected during growth of A. lipoferum on glucose; this species also possessed an NADP-linked glucose dehydrogenase and gluconokinase.     

Gluconate Catabolism in Rhizobium japonicum

Gluconate catabolism in Rhizobium japonicum ATCC 10324 was investigated by the radiorespirometric method and by assaying for key enzymes of the major energy-yielding pathways. Specifically labeled gluconate gave the following results for growing cells, with values expressed as per cent (14)CO(2) evolution: C-1 = 93%, C-2 = 57%, C-3 = 30%, C-4 = 70%, C-6 = 39%. The preferential release of (14)CO(2) from C-1 and C-4 indicate that gluconate is degraded primarily by the Entner-Doudoroff pathway but the inequalities between C-1 and C-4 and between C-3 and C-6 indicate that another pathway(s) also participates. The presence of gluconokinase and a system for converting 6-phosphogluconate to pyruvate also indicate a role for the Entner-Doudoroff pathway. The extraordinarily high yield of (14)CO(2) from C-1 labeled gluconate suggests that the other participating pathway is a C-1 decarboxylative pathway. The key enzyme of the pentose phosphate pathway, 6-phosphogluconate dehydrogenase, could not be demonstrated. Specifically labeled 2-ketogluconate and 2,5-diketogluconate were oxidized by gluconate grown cells and gave ratios of C-1 to C-6 of 2.73 and 2.61, respectively. These compare with a ratio of 2.39 obtained with specifically labeled gluconate. Gluconate dehydrogenase, the first enzyme in the ketogluconate pathway found in acetic acid bacteria, was found. Oxidation of specifically labeled pyruvate, acetate, succinate, and glutamate by gluconate-grown cells yielded the preferential rates of (14)CO(2) evolution expected from the operation of the tricarboxylic acid cycle. These data are consistent with the operation of the Entner-Doudoroff pathway and tricarboxylic acid cycle as the primary pathways of gluconate oxidation in R. japonicum. An ancillary pathway for the initial breakdown of gluconate would appear to be the ketogluconate pathway which enters the tricarboxylic acid cycle at alpha-ketoglutarate.     

Enzymes of Intermediary Carbohydrate Metabolism in the Obligate Autotrophs Thiobacillus thioparus and Thiobacillus neapolitanus

Levels of enzymes operative in the Embden-Meyerhof-Parnas (glycolytic) pathway, pentose phosphate cycle, citric acid cycle, and certain other phases of intermediary carbohydrate metabolism have been compared in Thiobacillus thioparus and T. neapolitanus. All enzymes of the glycolytic pathway except phosphofructokinase were demonstrated in both organisms. There were some striking quantitative differences between the two organisms with respect to the activities of the individual enzymes of the glycolytic pathway and the citric acid cycle. Qualitative differences were also found: the isocitrate dehydrogenase activity of T. thioparus is strictly nicotinamide adenine dinucleotide phosphate (NADP)-dependent, whereas that of T. neapolitanus is primarily nicotinamide adenine dinucleotide-dependent, activity with NADP being low; the glucose-6-phosphate dehydrogenase of T. thioparus is particulate, whereas that of T. neapolitanus is partly soluble and partly particulate; the 6-phosphogluconate dehydrogenase of T. thioparus is soluble, that of T. neapolitanus is partly soluble and partly particulate. All enzymes which function in the carbon reduction cycle were present at very high levels. In contrast, enzymes which operate exclusively in cycles other than the carbon reduction cycle were present at low levels. Of the enzymes not operative in the carbon reduction cycle that were examined, isocitric dehydrogenase had the highest specific activity. Both organisms possessed reduced nicotinamide adenine dinucleotide dehydrogenase activity. The qualitative and quantitative aspects of the data are discussed in relation to possible biochemical explanations of obligate autotrophy.     

A biochemical study of the intermediary carbon metabolism of Shewanella putrefaciens.

Cell extracts were used to determine the enzymes involved in the intermediary carbon metabolism of several strains of Shewanella putrefaciens. Enzymes of the Entner-Doudoroff pathway (6-phosphogluconate dehydratase and 2-keto-3-deoxy-6-phosphogluconate aldolase) were detected, but those of the Embden-Meyerhof-Parnas pathway were not. While several tricarboxylic acid cycle enzymes were present under both aerobic and anaerobic conditions, two key enzymes (2-oxoglutarate dehydrogenase and pyruvate dehydrogenase) were greatly diminished under anaerobic conditions. Extracts of cell grown anaerobically on formate as the sole source of carbon and energy were positive for hydroxypyruvate reductase, the key enzyme of the serine pathway in other methylotrophs, while no hexulose synthase activity was seen.     

The Deficient Carbohydrate Metabolic Pathways and the Incomplete Tricarboxylic Acid Cycle in an Obligately Autotrophic Hydrogen-oxidizing Bacterium

The activities of enzymes of carbohydrate metabolism, enzymes of the tricarboxylic acid cycle and some related enzymes were measured in cell-free extracts of strain TK-6, an extremely thermophilic, obligately autotrophic, aerobic hydrogen-oxidizing bacterium. Activities of phosphofructokinase, aldolase, pyruvate kinase, 6-phosphogluconate dehydrase and 2-keto-3-deoxy-6-phosphogluconate aldolase, key enzymes of the Embden-Meyerhof and the Entner-Doudoroff pathways were not found in the extracts. All of the tricarboxylic acid cycle enzymes except α-ketoglutarate dehydrogenase, and reduced nicotinamide adenine dinucleotide oxidase were present. These metabolic defects are considered to be one of the reasons for the obligate autotrophy of strain TK-6.     

Purification and Characterization of the Two 6-Phosphogluconate Dehydrogenase Species from Pseudomonas multivorans

The two species of 6-phosphogluconate dehydrogenase (EC 1.1.1.43) from Pseudomonas multivorans were resolved from extracts of gluconate-grown bacteria and purified to homogeneity. Each enzyme comprised between 0.1 and 0.2% of the total cellular protein. Separation of the two enzymes, one which is specific for nicotinamide adenine dinucleotide phosphate and the other which is active with nicotinamide adenine dinucleotide or nicotinamide adenine dinucleotide phosphate was facilitated by the marked difference in their respective isoelectric points, which were at pH 5.0 and 6.9. Comparison of the subunit compositions of the two enzymes indicated that they do not share common peptide chains. The enzyme active with nicotinamide adenine dinucleotide was composed of two subunits of about 40,000 molecular weight, and the nicotinamide adenine dinucleotide phosphate-specific enzyme was composed of two subunits of about 60,000 molecular weight. Immunological studies indicated that the two enzymes do not share common antigenic determinants. Reduced nicotinamide adenine dinucleotide phosphate strongly inhibited the 6-phosphogluconate dehydrogenase active with nicotinamide adenine dinucleotide by decreasing its affinity for 6-phosphogluconate. Guanosine-5'-triphosphate had a similar influence on the nicotinamide adenine dinucleotide phosphate-specific 6-phosphogluconate dehydrogenase. These results in conjunction with other data indicating that reduced nicotinamide adenine dinucleotide phosphate stimulates the conversion of 6-phosphogluconate to pyruvate by crude bacterial extracts suggest that in P. multivorans, the relative distribution of 6-phosphogluconate into the pentose phosphate and Entner-Doudoroff pathways might be determined by the intracellular concentrations of reduced nicotinamide adenine dinucleotide phosphate and purine nucleotides.     

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.     

Glucose Catabolism in Micrococcus sodonensis

The inability of Micrococcus sodonensis to grow on glucose as the sole source of carbon and energy was investigated. Estimation of pathways of glucose catabolism indicated that both the glycolytic and hexose monophosphate pathways are present in this organism. Comparative studies with Escherichia coli demonstrated that key enzymes for glucose catabolism were present in M. sodonensis in quantities equivalent to those of E. coli. The glucose-6-phosphate and 6-phosphogluconate dehydrogenases of M. sodonensis were nicotinamide adenine dinucleotide phosphate (NADP) specific, and glyceraldehyde-3-phosphate dehydrogenase was nicotinamide adenine dinucleotide specific. Transhydrogenase and reduced NADP oxidase were absent. Growth of the organism in the presence of glucose did not result in a repressed ability to oxidize tricarboxylic acid cycle intermediates, but these cells did have a decreased capacity for glucose degradation. The addition of substrates rich in growth-promoting substances, e.g., yeast extract, did not provide requisite nutrients for growth on glucose. Studies with (32)P suggest that M. sodonensis is incapable of synthesizing energy-rich phosphate compounds during the catabolism of glucose.     

Poly-β-hydroxybutyrate biosynthesis and the regulation of glucose metabolism in Azotobacter beijerinckii

Azotobacter beijerinckii possesses the enzymes of both the Entner-Doudoroff and the oxidative pentose phosphate cycle pathways of glucose catabolism and both pathways are subject to feedback inhibition by products of glucose oxidation. The allosteric glucose 6-phosphate dehydrogenase utilizes both NADP(+) and NAD(+) as electron acceptors and is inhibited by ATP, ADP, NADH and NADPH. 6-Phosphogluconate dehydrogenase (NADP-specific) is unaffected by adenosine nucleotides but is strongly inhibited by NADH and NADPH. The formation of pyruvate and glyceraldehyde 3-phosphate from 6-phosphogluconate by the action of the Entner-Doudoroff enzymes is inhibited by ATP, citrate, isocitrate and cis-aconitate. Glyceraldehyde 3-phosphate dehydrogenase is unaffected by adenosine and nicotinamide nucleotides but the enzyme is non-specific with respect to NADP and NAD. Citrate synthase is strongly inhibited by NADH and the inhibition is reversed by the addition of AMP. Isocitrate dehydrogenase, a highly active NADP-specific enzyme, is inhibited by NADPH, NADH, ATP and by high concentrations of NADP(+). These findings are discussed in relation to the massive synthesis of poly-beta-hydroxybutyrate that occurs under certain nutritional conditions. We propose that synthesis of this reserve material, to the extent of 70% of the dry weight of the organism, serves as an electron and carbon ;sink' when conditions prevail that would otherwise inhibit nitrogen fixation and growth.     

Cellobiose catabolism in the haloalkaliphilic hydrolytic bacterium <Emphasis Type="Italic">Alkaliflexus imshenetskii</Emphasis>

Cellobiose metabolism was studied in Alkaliflexus imshenetskii, a haloalkaliphilic hydrolytic bacterium capable of utilizing certain polymers of plant origin, as well as mono- and disaccharides. The major products of cellobiose fermentation by the bacterium were succinate and acetate, and formate was a minor product. Cellobiose could be split into glucose molecules by both β-glucosidase (hydrolytic pathway) and phosphorylase (phosphorolytic pathway); the activity of the former enzyme was two orders of magnitude higher (3600 nmol/(min mg) versus 36 nmol/(min mg)). In cell extracts of the bacterium, high activities of the Embden-Meyerhof-Parnas pathway enzymes—hexokinase, glucose-phosphate isomerase, and phosphofructokinase—were revealed, as well as the activities of glucose-6-phosphate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, and key enzymes of the Entner-Doudoroff pathway—6-phospho-gluconate dehydratase and 2-keto-3-deoxy-6-phospho-gluconate aldolase. Neither the activity of the key enzyme of the hexose-mono-phosphate pathway, 6-phospho-gluconate dehydrogenase, nor the activities of the key enzymes of the modified Entner-Doudoroff pathway, glucose dehydrogenase and 2-keto-3-deoxy-gluconate kinase, were revealed.