Leishmania mexicana: enzyme activities of amastigotes and promastigotes and their inhibition by antimonials and arsenicals

A major difference between the metabolism of Leishmania species amastigotes and cultured promastigotes was found in the area of CO2 fixation and phosphoenolpyruvate metabolism. Malate dehydrogenase (EC 1.1.1.37) and phosphoenolpyruvate carboxykinase (EC 4.1.1.49) were at much higher activities in amastigotes than promastigotes of both L. m. mexicana and L. donovani, whereas the reverse was true of pyruvate kinase (EC 2.7.1.40). Pyruvate carboxylase (EC 6.4.1.1) and malic enzyme (carboxylating) (EC 1.1.1.40) could not be detected in L. m. mexicana amastigotes. Promastigotes of L. m. mexicana had a high NAD-linked glutamate dehydrogenase activity in comparison to amastigotes, whereas NADP-linked glutamate dehydrogenase activity was detected only in amastigotes. Leishmania m. mexicana culture promastigotes were killed in vitro by the trivalent antimonial Triostam (LD50, 20 micrograms/ml) and the trivalent arsenical melarsen oxide (LD50, 20 micrograms/ml), but they were unaffected by Pentostam. Neither antimonial drug significantly inhibited leishmanial hexokinase (EC 2.7.1.2), phosphofructokinase (EC 2.7.1.11), pyruvate kinase, malate dehydrogenase or phosphoenolpyruvate carboxykinase, whereas melarsen oxide was a potent inhibitor of all the enzymes tested except phosphoenolpyruvate carboxykinase.     

On the cellular localization of phosphoenolpyruvate carboxylase in Sorghum leaves

The localization of phosphoenol pyruvate carboxylase (EC 4.1.1.3.1.) in the leaf cells of Sorghum vulgare was investigated by using three techniques: the conventional aqueous and non aqueous methods gave conflicting results; the immunocytochemical techniques clearly showed that the enzyme is predominantly located in the cytoplasm of mesophyll cells.Abbreviations PEP phosphoenol pyruvate - PAG polyacrylamide gel - NADP MDH NADP malate dehydrogenase - FITC fluorescein isothiocyanate - SAB serum albumine bovine - DTT dithiothreitol - MDH malate dehydrogenase - ME malic enzyme - PBS phosphate buffer saline - PAP peroxidase anti-peroxidase     

Respiration-associated components of Mollicutes.

No cytochrome pigments were detected by difference (reduced minus oxidized) spectroscopy at liquid nitrogen temperature in whole-cell preparations or membrane fractions of Acholeplasma axanthum S273, Acholeplasma equifetale N93, Acholeplasma granularum BTS39, Acholeplasma laidlawii B-PG9, Acholeplasma modicum PG-49, Acholeplasma oculi 19L, Mycoplasma arginini G230, Mycoplasma arthritidis 07, Mycoplasma pneumoniae FH, and Mycoplasma pulmonis JB. All ten Mollicutes species examined contained iron of unknown function (3.0 to 15.3 nmol of iron per mg of protein). Relatively small amounts of acid-labile sulfide were found in all fractions (0.10 to 1.07 nmol of acid-labile sulfide per mg of protein). The data suggest that, as Mollicutes lack cytochrome pigments, they would synthesize most if not all adenosine triphosphate at the substrate level.     

Activity of enzymes of carbon metabolism during the induction of Crassulacean acid metabolism in Mesembryanthemum crystallinum L.

The maximum extractable activities of twenty-one photosynthetic and glycolytic enzymes were measured in mature leaves of Mesembryanthemum crystallinum plants, grown under a 12 h light 12 h dark photoperiod, exhibiting photosynthetic characteristics of either a C₃ or a Crassulacean acid metabolism (CAM) plant. Following the change from C₃ photosynthesis to CAM in response to an increase in the salinity of in the rooting medium from 100 mM to 400 mM NaCl, the activity of phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) increased about 45-fold and the activities of NADP malic enzyme (EC 1.1.1.40) and NAD malic enzyme (EC 1.1.1.38) increased about 4- to 10-fold. Pyruvate, Pi dikinase (EC 2.7.9.1) was not detected in the non-CAM tissue but was present in the CAM tissue; PEP carboxykinase (EC 4.1.1.32) was detected in neither tissue. The induction of CAM was also accompanied by large increases in the activities of the glycolytic enzymes enolase (EC 4.2.1.11), phosphoglyceromutase (EC 2.7.5.3), phosphoglycerate kinase (EC 2.7.2.3), NAD glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12), and glucosephosphate isomerase (EC 2.6.1.2). There were 1.5- to 2-fold increases in the activities of NAD malate dehydrogenase (EC 1.1.1.37), alanine and aspartate aminotransferases (EC 2.6.1.2 and 2.6.1.1 respectively) and NADP glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.13). The activities of ribulose-1,5-bisphosphate (RuBP) carboxylase (EC 4.1.1.39), fructose-1,6-bisphosphatase (EC 3.1.3.11), phosphofructokinase (EC 2.7.1.11), hexokinase (EC 2.7.1.2) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) remained relatively constant. NADP malate dehydrogenase (EC 1.1.1.82) activity exhibited two pH optima in the non-CAM tissue, one at pH 6.0 and a second at pH 8.0. The activity at pH 8.0 increased as CAM was induced. With the exceptions of hexokinase and glucose-6-phosphate dehydrogenase, the activities of all enzymes examined in extracts from M. crystallinum exhibiting CAM were equal to, or greater than, those required to sustain the maximum rates of carbon flow during acidification and deacidification observed in vivo. There was no day-night variation in the maximum extractable activities of phosphoenolpyruvate carboxylase, NADP malic enzyme, NAD malic enzyme, fructose-1,6-bisphosphatase and NADP malate dehydrogenase in leaves of M. crystallinum undergoing CAM.Abbreviations CAM Crassulacean acid metabolism - PEP phosphoenolpyruvate - RuBP ribulose-1,5-bisphosphate     

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     

Activities of enzymes concerned with pyruvate and oxaloacetate metabolism in the heart and liver of developing sheep.

1. In order to assess whether the potential ability of heart ventricular muscle and liver to metabolise substrates such as alanine, aspartate and lactate varies as the sheep matures and its nutrition changes, the activities of the following enzymes were determined in tissues of lambs obtained at varying intervals between 50 days after conception to 16 weeks after birth and in livers from adult pregnant ewes: lactate dehydrogenase (EC 1.1.1.27), alanine aminotransferase (EC 2.6.1.2), pyruvate kinase (EC 2.7.1.40), pyruvate carboxylase (EC 6.4.1.1), phosphoenolpyruvate carboxykinase (GTP)(EC 4.1.1.32), malate dehydrogenase (EC 1.1.1.37), aspartate aminotransferase (EC 2.6.1.1) and citrate (si)-synthase (EC 4.1.3.7). 2. In the heart a most marked increase in alanine aminotransferase activity was found throughout development. During this period the activities of citrate (si)-synthase, lactate dehydrogenase and pyruvate carboxylase also increased. There were no substantial changes in the activities of aspartate aminotransferase, malate dehydrogenase or pyruvate kinase. Pyruvate kinase activities were five times greater in the heart compared with those found in the liver. No significant activity of phosphoenolpyruvate carboxykinase (GTP) was detected in heart muscle. 3. In the liver the activities of both alanine aminotransferase and aspartate aminotransferase increased immediately following birth although the activity of alanine aminotransferase was lower in livers of pregnant ewes than in any of the lambs. As with alanine aminotransferase the highest activities of lactate dehydrogenase were found during the period of postnatal growth. No marked changes were observed in malate dehydrogenase or citrate (si)-synthase activities during development. A small decline in pyruvate kinase activity occurred whilst the activities of pyruvate carboxylase and phosphoenolpyruvate carboxykinase (GTP) tended to rise during development.     

Interactions of Phosphoenolpyruvate Carboxylase and Pyruvic Kinase in Developing Soybean Seeds

Developing soybean seeds contain phosphoenolpyruvate (PEP) carboxylase,pyruvic kinase, malate dehydrogenase, aspartate aminotransferase,alanine aminotransferase and malic enzyme activities. PEP carboxylasemay be important in competing with pyruvic kinase and directinga portion of glycolytic carbon towards oxaloacetate synthesis.The oxaloacetate can then be converted to aspartate and malate.Malic enzyme produces pyruvate and NADPH from malate, and thismay be an important additional source of reducing power forlipid biosynthesis. In the presence of high levels of PEP carboxylaseit is possible to demonstrate PEP formation by pyruvic kinase.PEP carboxylase and pyruvic kinase independently compete forPEP in a mixed system. Soybean seed extracts readily convertedradioactive PEP into alanine and aspartate when supplementedwith ADP, Mg²⁺, K+, HCO₃– and glutamate. Under varyingconditions of pH, metal ions, PEP, enzyme concentration andtime both alanine and aspartate were always produced. Possiblythe final products of glycolysis should be considered as pyruvateand oxaloacetate in plants. (Received April 22, 1981; Accepted June 26, 1981)     

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)     

Immunocytochemical study of phosphoenolpyruvate carboxylase in nodulated Alnus glutinosa

The activities of phosphoenolpyruvate carboxylase (PEP carboxylase, EC 4.1.1.3.1) have been investigated in various organs of young nodulated Alnus glutinosa. The root nodules exhibited the highest specific enzyme activity when compared with the one in roots and leaves. Furthermore, in the root nodules the PEP carboxylase was predominantly localized in the cytosol of the large cortical cells containing the endophyte vesicles.Abbreviations PEP carboxylase phosphoenolpyruvate carboxylase - MDH malate dehydrogenase - PVP polyvinylpyrrolidone - PBS phosphate buffer saline     

Enzymic changes in rabbit and rat mammary gland during the lactation cycle

1. Activities of glucose 6-phosphate dehydrogenase (EC 1.1.1.49), 6-phosphogluconate dehydrogenase (EC 1.1.1.44), isocitrate dehydrogenase (EC 1.1.1.42), malate dehydrogenase (EC 1.1.1.37), malate dehydrogenase (decarboxylating) (EC 1.1.1.40), and pyruvate carboxylase (EC 6.4.1.1) were determined in subcellular fractions of mammary gland from rabbits during pregnancy, at different stages of lactation and during weaning. The results were compared with those obtained in similar experiments with rat mammary gland. 2. Three bases of expression of the activity of enzymes in the particle-free supernatant fraction of mammary gland were compared. During lactation, activity expressed per mg. of particle-free supernatant protein (uncorrected for milk protein) correlated well with that expressed per mug. of DNA phosphorus. The disadvantages of expressing activities per g. wet wt. are discussed. 3. The major differences between the two tissues were: (a) neither malate dehydrogenase (decarboxylating) nor a soluble form of pyruvate carboxylase could be detected in rabbit mammary gland at any stage of the lactation cycle; (b) isocitrate dehydrogenase increased in activity during lactation in rabbit mammary gland, but not in that of the rat. 4. Pyruvate carboxylase in the mitochondrial fraction of rabbit mammary gland, and in both the mitochondrial and the soluble fractions of rat mammary gland, did not change in activity during lactation. 5. For each tissue, the NADP-dependent dehydrogenases studied had a high activity at all stages of the lactation cycle compared with the rate of fatty acid synthesis at mid-lactation. The significance of these results is discussed with respect to the supply of NADPH via NADH.     

Some enzyme activities associated with the chlorophyll containing layers of the immature barley pericarp

Summary Some photosynthetic and biochemical properties of the chlorophyl containing layers of the pericarp of developing barley have been investigated. The tissue changes from pale green to bright green early in development, chlorophyll disappearing only at the later stages of maturity. It contains chloroplasts and probably amyloplasts and starch bearing chloroplasts. It is capable of high rates of light dependent oxygen evolution. It has been shown that the enzyme phosphoenol pyruvate carboxylase (EC 4.1.1.31) is present in the pericarp and is 100 times as active in carbon dioxide fixation as ribulose diphosphate carboxylase (EC 4.1.1.39). Other enzymes present in the pericarp are phosphoenol pyruvate synthetase, pyrophosphatase (EC 3.6.1.1), malate NAD and NADP dehydrogenases (EC 1.1.1.37), malic enzyme (EC 1.1.1.40), and fructose 1,6 diphosphatase (EC 3.1.3.11).Abbreviations RDP Ribulose 1,5-diphosphate - PEP phosphoenol pyruvate     

Correlations between photosynthetic enzymes,CO2-fixation and plastid structure in an albino mutant of Zea mays L.

Summary An albino seedling of Zea mays L. was investigated for its potential for CO₂-assimilation. In the mesophyll the number, dimensions and fine structure of chloroplasts are drastically reduced but to a lesser extent in the bundle sheath. Chlorophyll concentration is zero and carotenoid concentration almost zero. Albinism also exerts a strong influence on the stroma of bundle sheath chloroplasts; ribulose-1.5-biphosphate carboxylase (EC 4.1.1.39) activity and glyceraldehyde-3-phosphate dehydrogenase (NADP) (EC 1.2.1.13) activity is not detectable. The C₄-enzymes phosphoenolpyruvate carboxylase (EC 4.1.1.31) and malate dehydrogenase (decarboxylating) (EC 1.1.1.40) and the non-photosynthetic linked enzymes malate dehydrogenase (NAD) (EC 1.1.1.37), aspartate-2-oxoglutarate aminotransferase (EC 1.1.1.37), aspartate-2-oxoglutarate aminotransferase (EC 2.6.1.1.) and glyceraldehyde-3-phosphate dehydrogenase (NAD) (EC 1.2.1.1.) are present in the albino seedling with activities comparable to those in etiolated maize seedlings. The potential for CO₂ fixation of the albino seedlings exceeds that of comparable dark seedlings considerably. The results are discussed with regard to enzyme localization of the C₄ pathway of photosynthesis.Abbreviations Aspartate aminotransferase L-aspartate-2-oxoglutarate aminotransferase-EC 2.6.1.1. - GAPDH (NAD) glyceraldehyde-3-phosphate dehydrogenase (NAD dep.)-EC 1.2.1.12 - GAPDH (NADP) glyceraldehyde-3-phosphate dehydrogenase (NADP dep.)-EC 1.2.1.13 - malic enzyme malate dehydrogenase (NADP dep., decarboxylating)-EC 1.1.1.40 - MDH malate dehydrogenase (NAD dep.)-1.1.1.37 - PEP carboxylase phosphoenolpyruvate carboxylase-EC 4.1.1.31 - RuDP carboxylase ribulose-1.5-biphosphate carboxylase-EC 4.1.1.39     

Activities of oxidative enzymes in mycoplasmas.

The activities of several oxidoreductases were measured in three fermentative and two nonfermentative Mycoplasma species that were grown under aerobic or anaerobic conditions. Acholeplasma laidlawii MG, Mycoplasma hyorhinis GDL, and Mycoplasma pneumoniae FH had very high apparent activities of pyruvate dehydrogenase and pyruvate dehydrogenase complex compared with the activities of mammalian fibroblasts or human platelet-enriched preparations, while Mycoplasma salivarium VV and Mycoplasma arthritidis 07 had very low apparent activities of these two enzymes. Strictly anaerobic growth diminished both enzymatic activities. The activity of alpha-ketoglutarate dehydrogenase complex was minimal in all five mycoplasmas that were grown under aerobic conditions, anaerobic conditions, or both. All the mycoplasmas that were examined exhibited lactate dehydrogenase and NADH-dichlorophenol indophenol oxidoreductase activities. The properties of mycoplasmal pyruvate dehydrogenase complex suggest that it differs from the mammalian enzyme.     

The metabolic pathways of Acholeplasma and Mycoplasma: an overview

The metabolism of the Mollicutes Acholeplasma and Mycoplasma may be characterized as restricted, for example, by virtue of the apparent absence of cytochrome pigments. Some Mollicutes have lowered ECA values during their logarithmic growth phase, which we speculate may be related to insufficient substrate phosphorylation or insufficient ATP synthesis linked to glycolysis. We found that PEP is carboxylated by preparations of A. laidlawii, but not by other Mollicutes; thus in this organism oxaloacetate from PEP may be a link to other pathways. We found phosphoribosylpyrophosphate in A. laidlawii, which suggests that ribosylation of purines and pyrimidines occurs in Mollicutes other than M. mycoides.     

Junctions of catabolic and anabolic pathways in Zymomonas mobilis: phosphoenolpyruvate carboxylase and malic enzyme

Summary The synthesis of oxalacetate and malate in the ethanol-producing bacterium Zymomonas mobilis have been investigated. Cell-free extracts were examined for pyruvate carboxylase, phosphoenolpyruvate (PEP) carboxylase, PEP carboxytransphosphorylase, PEP carboxykinase, and malic enzyme, but only PEP carboxylase and nicotine adenine dinucleotide (NAD)-dependent malic enzyme activities could be detected. The PEP carboxylase, partially purified from extracts, was not affected by acetyl-coenzyme A. Intermediates of the tricarboxylic acid cycle and aspartate inhibited the enzyme competitively with PEP. Of these, citrate and -ketoglutarate were the strongest inhibitors. The physiological roles of PEP carboxylase and malic enzyme in Z. mobilis are discussed.Dedicated to Prof. Dr. A. Fiechter, ETH Zürich, on the occasion of his 65th birthday     

Inhibition of phosphoenolpyruvate carboxylase by malate

Malate has been noted to be a `mixed' inhibitor of phosphoenolpyruvate (PEP) carboxylase. The competitive portion of this inhibition appears to be fairly constant regardless of the condition of the enzyme being measured, but the noncompetitive (V-type) inhibition is subject to variation depending on the source of the enzyme, its storage condition, the presence or absence of various ligands, and differences in pH. In the case of the maize (Zea mays L.) phosphoenolpyruvate carboxylase (PEPC), the V-type inhibition by malate is much less pronounced at pH 8 than at pH 7. Examination of the response of the maize PEPC to PEP concentration reveals a pronounced cooperativity at pH 8 which is not present at pH 7, and which results in the disappearance of the V-type inhibition at pH 8. The ability of high concentrations of PEP to convert PEPC from a form readily inhibited by malate to one resistant to malate inhibition has been previously demonstrated and we attribute the cooperativity shown at pH 8 to this response to high levels of PEP. Support for this proposal is provided by studies of the enzyme at pH 7 and pH 8 run in 20% glycerol. In this case there was no V-type inhibition of PEPC at either pH. Treatment with 20% glycerol has been shown to result in the aggregation of maize PEPC.     

Effects of fusicoccin on the activity of a key pH-stat enzyme,PEP-carboxylase

The phytotoxin fusicoccin (FC) causes rapid synthesis of malate in coleoptile tissues, presumably via phosphoenolpyruvate (PEP) carboxylase coupled with malate dehydrogenase. The possibility that FC directly affects PEP carboxylase in Avena sativa L. and Zea mays L. coleoptiles was studied and rejected. The activity of this enzyme is unaffected by FC whether FC is added in vitro or a pretreatment to the live material. FC does not change the sensitivity of the enzyme to bicarbonate or malate. The activity of FC, instead, appears to be indirect. The pH sensitivity of PEP carboxylase is such that its activity, and thus the rate of malate synthesis, may be enhanced by an increase in cytoplasmic pH accompanying FC-induced H⁺ excretion. Since the enzyme is also particularily sensitive to bicarbonate levels, malate synthesis may also be enhanced by FC-induced uptake or generation of CO₂.     

Membrane enzymes associated with the dissimilation of some citric acid cycle substrates and production of extracellular oxidation products in chemostat cultures of Pseudomonas fluorescens

Enzyme activities forming extracellular products from succinate, fumarate, and malate were examined using washed cell suspensions of Pseudomonas fluorescens from chemostat cultures. Membrane-associated enzyme activities (glucose, gluconate, and malate dehydrogenases), producing large accumulations of extracellular oxidation products in carbon-excess environments, have previously been found in P. fluorescens. Investigations carried out here have demonstrated the presence in this microorganism of a malic enzyme activity which produces extracellular pyruvate from malate in carbon-excess environments. Although the three membrane dehydrogenase enzymes decrease significantly in carbon-limited chemostat cultures, malic enzyme activity was found to increase fourfold under these conditions. The regulation of malate dehydrogenase and malic enzyme by malate or succinate was similar. Malate dehydrogenase increased and malic enzyme decreased in carbon-excess cultures. The opposite effect was observed in carbon-limited cultures. When pyruvate or glucose was used as the carbon source, malate dehydrogenase was regulated similarly by the available carbon concentration, but malic enzyme activity producing extracellular pyruvate was not detected. While large accumulations of extracellular oxalacetate and pyruvate were produced in malate-excess cultures, no extracellular oxidation products were detected in succinate-excess cultures. This may be explained by the lack of detectable activity for the conversion of added external succinate to extracellular fumarate and malate in cells from carbon-excess cultures. In cells from carbon-limited (malate or succinate) cultures, very active enzymes for the conversion of succinate to extracellular fumarate and malate were detected. Washed cell suspensions from these carbon-limited cultures rapidly oxidized added succinate to extracellular pyruvate through the sequential action of succinate dehydrogenase, fumarase, and malic enzyme. Succinate dehydrogenase and fumarase activities producing extracellular products were not detected in cells from chemostat cultures using pyruvate or glucose as the carbon source. Uptake activities for succinate, malate, and pyruvate also were found to increase in carbon-limited (malate or succinate) and decrease in carbon-excess cultures. The role of the membrane-associated enzymes forming different pathways for carbon dissimilation in both carbon-limited and carbon-excess environments is discussed.     

Malate Dehydrogenase Mutants in Escherichia coli K-12

Mutants devoid of malate dehydrogenase activity have been isolated in Escherichia coli K-12. They do not possess detectable malate dehydrogenase when grown aerobically or anaerobically on glucose as sole carbon source. All mutants revert spontaneously; a few partial revertants have been found with a malate dehydrogenase exhibiting altered electrophoretic mobility. Therefore, only one such enzyme appears to exist in the strains examined. No evidence could be obtained for the presence of a malate dehydrogenase not linked to nicotinamide adenine dinucleotide. Mutants deficient in both malate dehydrogenase and phosphoenol pyruvate carboxylase activities will grow anaerobically on minimal glucose plus succinate medium; also, malate dehydrogenase mutants do not require succinate for anaerobic growth on glucose. The anaerobic pathway oxaloacetate to succinate or succinate to aspartate appears to be accomplished by aspartase. Malate dehydrogenase is coded for by a locus somewhere relatively near the histidine operon, i.e., a different chromosomal location than that known for other citric acid cycle enzymes.