Glycolytic enzyme interactions with tubulin and microtubules

Interactions of the glycolytic enzymes glucose-6-phosphate isomerase, aldolase, glyceraldehyde-3-phosphate dehydrogenase, triose-phosphate isomerase, enolase, phosphoglycerate mutase, phosphoglycerate kinase, pyruvate kinase, lactate dehydrogenase type-M, and lactate dehydrogenase type-H with tubulin and microtubules were studied. Lactate dehydrogenase type-M, pyruvate kinase, glyceraldehyde-3-phosphate dehydrogenase, and aldolase demonstrated the greatest amount of co-pelleting with microtubules. The presence of 7% poly(ethylene glycol) increased co-pelleting of the latter four enzymes and two other enzymes, glucose-6-phosphate isomerase, and phosphoglycerate kinase with microtubules. Interactions also were characterized by fluorescence anisotropy. Since the KD values of glyceraldehyde-3-phosphate dehydrogenase, pyruvate kinase and lactate dehydrogenase for tubulin and microtubules were all found to be between 1 and 4 microM, which is in the range of enzyme concentration in cells, these enzymes are probably bound to microtubules in vivo. These observations indicate that interactions of cytosolic proteins, such as the glycolytic enzymes, with cytoskeletal components, such as microtubules, may play a structural role in the formation of the microtrabecular lattice.     

Degradation of glucose-metabolizing enzymes in the rat small intestine during starvation

1. The degradation rates and half-lives of hexokinase, 6-phosphogluconate dehydrogenase, lactate dehydrogenase, pyruvate kinase, glucose 6-phosphate dehydrogenase, phosphoglycerate kinase and aldolase were calculated from measurements of the decline in activities of these enzymes in rat small intestine during starvation. 2. The half-lives of the enzymes are: hexokinase, 5.7h; 6-phosphogluconate dehydrogenase, 7.6h; glucose 6-phosphate dehydrogenase, 6.0h; pyruvate kinase, 8.9h; lactate dehydrogenase, 8.7h; phosphoglycerate kinase, 8.7h; aldolase, 5.1h. 3. The significance of the results is discussed with respect to the regulation of enzyme concentrations in response to changes in diet.     

Glycolytic enzyme levels in synaptosomes

The specific activities of glucosephosphate isomerase, aldolase, triosephosphate isomerase, glyceraldehydephosphate dehydrogenase, phosphoglycerate kinase, phosphoglycerate mutase, pyruvate kinase and lactate dehydrogenase were all higher in the synaptoplasmic fraction from rat brain than in 100,000 g supernatant fraction of rat brain homogenates when the supernatants were prepared in high ionic strength solutions. Four enzymes in synaptosomes and two enzymes in homogenates were associated with particulate fractions as indicated by the large increase in specific activity of the enzymes when samples were treated with 0.3 M KCl before centrifugation. Glucosephosphate isomerase, aldolase, pyruvate kinase and lactate dehydrogenase were the enzymes that showed a large increase in specific activity following salt treatment of isolated, synaptosomal membrane while aldolase and pyruvate kinase were the two enzymes which showed a large increase in specific activity in the high speed supernatant fractions. Because the specific activities of many enzymes are found to be elevated not only in synaptosomes but in synaptosomal membrane fractions it is suggested that these enzymes may provide the potential for significantly enhanced glycolysis at these locations.     

Enzyme activities of human erythrocyte ghosts: effects of various treatments

Summary Ghosts of human erythrocytes prepared by hypotonic hemolysis were assayed for aldolase, triosephosphate isomerase, glyceraldehyde phosphate dehydrogenase, phosphoglycerate kinase, pyruvate kinase, lactate dehydrogenase, and glutathione peroxidase and reductase. Cryptic activity of the enzymes was demonstrated by an increase in activity on dilution with water, which caused fragmentation of the ghosts. Aldolase and glyceraldehyde phosphate dehydrogenase were classed as firmly bound; phosphoglycerate kinase was intermediate; the others were loosely bound. Triton X-100 increased the activities of aldolase, glyceraldehyde phosphate dehydrogenase, and phosphoglycerate kinase. The pH of the medium had little effect upon the firmly bound enzymes but it markedly affected the retention of hemoglobin and the activities of the loosely bound enzymes. The presence of Mg or Ca ions enhanced the retention of hemoglobin and the activity of lactate dehydrogenase and pyruvate kinase, with little effect on aldolase and glyceraldehyde phosphate dehydrogenase. Ghosts diluted in water disintegrated into fragments and tubules or vesicles; Mg or Ca at 1mm afforded protection against this. When ghosts were treated with Triton X-100 and adenosine triphosphate, they contracted to about one-seventh of their volume. The shrunken ghosts had lost a considerable proportion of their cholesterol and protein to the medium.     

Organization of enzymes of glycolysis and of glutathione metabolism in human red cell membranes.

1) The activities of 16 enzymes of glycolysis and of glutathione metabolism were determined in intact human red cell membranes (ghosts) which were prepared by hypotonic hemolysis. 2) Enzymes and hemoglobin of the ghosts were resolved by two toluene extractions. Only the four enzymes hexokinase, fructose-bisphosphate aldolase, glyceraldehyde-phosphate dehydrogenase and pyruvate kinase could not be released completely from the ghosts. 3) The residual membrane fraction, which was obtained after the toluene extraction of ghosts prepared at 30 imOsM, contained 0.02% of the original hemoglobin content of the red cell. Between 6.5 and 23% of the hemolysate activities of glyceraldehyde-phosphate dehydrogenase, phosphoglycerate kinase, pyruvate kinase and fructose-bisphosphate aldolase were detected in this fraction after mechanical disruption. 4) Sonication of intact ghosts increased the activities of fructose-bisphosphate aldolase, pyruvate kinase and phosphoglycerate kinase. 5) In "white" ghosts prepared at 5 imOsM phosphate buffer which contained 0.5% of the original hemoglobin the activities of fructose-bisphosphate aldolase and glyceraldehyde-phosphate dehydrogenase were detected at high levels. The activities of pyruvate kinase and phosphoglycerate kinase were low in these preparations. 6) The results indicate that one part of all enzymes is loosely attached to the inner surface of the membrane as is hemoglobin. A second part, the "cryptic enzyme activity", is available after resolving by toluene. A residual part of four enzymes is firmly bound to the membrane. Two of them (fructose-bisphosphate aldolase and glyceraldehyde-phosphate dehydrogenase) are oriented toward the inner surface of the membrane, whereas pyruvate kinase and phosphoglycerate kinase are hidden in the lipid core of the membrane.     

Activity and androgenic control of glycolytic enzymes in the epididymis and epididymal spermatozoa of the rat.

1. Procedures were developed for the extraction and assay of glycolytic enzymes from the epididymis and epididymal spermatozoa of the rat. 2. The epididymis was separated into four segments for analysis. When rendered free of spermatozoa by efferent duct ligation, regional differences in enzyme activity were apparent. Phosphofructokinase, glycerol phosphate dehydrogenase and glucose 6-phosphate dehydrogenase were more active in the proximal regions of the epididymis, whereas hexokinase, lactate dehydrogenase and phosphorylase were more active in the distal segment. These enzymes were less active in the epididymis of castrated animals and less difference was apparent between the proximal and distal segments. However, the corpus epididymidis from castrated rats had lower activities of almost all enzymes compared with other epididymal segments. 3. Spermatozoa required sonication to obtain satisfactory enzyme release. Glycolytic enzymes were more active in spermatozoa than in epididymal tissue, being more than 10 times as active in the case of hexokinase, phosphoglycerate kinase and phosphoglycerate mutase. 4. The specific activities of a number of enzymes in the epididymis were dependent on the androgen status of the animal. These included hexokinase, phosphofructokinase, aldolase, glyceraldehyde phosphate dehydrogenase, phosphoglycerate kinase, pyruvate kinase, glycerol phosphate dehydrogenase, glucose 6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase and phosphorylase. 5. The caput and cauda epididymidis differed in the extent to which enzyme activities changed in response to an altered androgen status. The most notable examples were hexokinase, phosphofructokinase, aldolase, phosphoglycerate kinase, 6-phosphogluconate dehydrogenase and phosphorylase.     

Metabolism of various carbon sources by Azospirillum brasilense

Azospirillum brasilense Sp7 and two mutants were examined for 19 carbon metabolism enzymes. The results indicate that this nitrogen fixer uses the Entner-Doudoroff pathway for gluconate dissimilation, lacks a catabolic but has an anabolic Embden-Meyerhof-Parnas hexosephosphate pathway, has amphibolic triosephosphate enzymes, lacks a hexose monophosphate shunt, and has lactate dehydrogenase, malate dehydrogenase, and glycerokinase. The mutants are severely deficient in phosphoglycerate and pyruvate kinase and also have somewhat reduced levels of other carbon enzymes.     

Key enzymes of carbohydrate metabolism as targets of the 11.5-kDa Zn(2+)-binding protein (parathymosin)

The 11.5-kDa Zn(2+)-binding protein (ZnBP) was covalently linked to Sepharose. Affinity chromatography with a cytosolic subfraction from liver resulted in purification of a predominant 38-kDa protein. In comparable experiments with brain cytosol a 39-kDa protein was enriched. The ZnBP-protein interactions were zinc-specific. Both proteins were identified as fructose-1,6-bisphosphate aldolase. Experiments with crude cytosol showed zinc-specific interaction of additional enzymes involved in carbohydrate metabolism. From liver cytosol greater than 90% of the following enzymes were specifically retained: aldolase, phosphofructokinase-1, hexokinase/glucokinase, glucose-6-phosphate dehydrogenase, glycerol-3-phosphate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, and fructose-1,6-bisphosphatase. Glucose-6-phosphate isomerase, phosphoglycerate kinase, enolase, lactate dehydrogenase, and most of triosephosphate isomerase remained unbound. From L-type pyruvate kinase only the phosphorylated form seems to interact with ZnBP. Using brain cytosol hexokinase, phosphofructokinase-1, and aldolase were completely bound to the affinity column, whereas glucose-6-phosphate isomerase, phosphoglycerate kinase, enolase, lactate dehydrogenase, pyruvate kinase, and most of triose-phosphate isomerase remained unbound. The behavior of glucose-6-phosphate dehydrogenase and glycerol-3-phosphate dehydrogenase from this tissue could not be followed. A possible function of ZnBP in supramolecular organization of carbohydrate metabolism is proposed.     

The effect of dietary and hormonal conditions on the activities of glycolytic enzymes in rat epididymal adipose tissue

1. Measurements were made of the activities of nine glycolytic enzymes in epididymal adipose tissues obtained from rats that had undergone one of the following treatments: starvation; starvation followed by re-feeding with bread or high-fat diet; feeding with fat without preliminary starvation; alloxan-diabetes; alloxan-diabetes followed by insulin therapy. 2. In general, the activities of the glycolytic enzymes of adipose tissue, unlike those of liver, were not greatly affected by the above treatments. 3. The ;key' glycolytic enzymes, phosphofructokinase and pyruvate kinase, were generally no more adaptive in response to physiological factors than other glycolytic enzymes such as glucose phosphate isomerase, fructose diphosphate aldolase, triose phosphate isomerase, glycerol 3-phosphate dehydrogenase, phosphoglycerate kinase and lactate dehydrogenase. 4. Adiposetissue pyruvate kinase did not respond to feeding with fat in a manner similar to the liver enzyme. 5. Glyceraldehyde phosphate dehydrogenase had a behaviour pattern unlike the other eight glycolytic enzymes studied in that its activity was depressed by feeding with fat and was not restored to normal by re-feeding with a high-fat diet after starvation. These results are discussed in relation to the requirements of adipose tissue for glycerol phosphate in the esterification of fatty acids. 6. A statistical analysis of the results permitted the writing of linear equations describing the relationships between the activities of eight of the enzymes studied. 7. Evidence is presented for the existence of two constant-proportion groups amongst the enzymes studied, namely (i) glucose phosphate isomerase, phosphoglycerate kinase and lactate dehydrogenase, and (ii) triose phosphate isomerase, fructose diphosphate aldolase and pyruvate kinase. 8. Mechanisms for maintaining the observed relationships between the activities of the enzymes in the tissue are discussed.     

Trypanosoma brucei: activities and subcellular distribution of glycolytic enzymes from differently disrupted cells

The effect of disruption procedure on the subcellular distribution and the activities of 11 enzymes catalyzing the glycolytic pathway in Trypanosoma brucei has been studied. The activities of the enzymes varied with the lytic procedure used. Maximum specific enzyme activity values were obtained after treatment with saponin whereas digitonin treatment gave the lowest results. The intracellular location of the enzymes was examined by means of differential centrifugation following cell lysis with saponin, Triton X-100, digitonin, or by freezing and thawing. Irrespective of the method of cell lysis employed, the six enzymes, hexokinase, phosphofructokinase, aldolase, phosphoglycerate kinase, glycerol phosphate dehydrogenase, and glycerokinase, were particulate. Of the remaining 5 enzymes, digitonin liberates only phosphoglycerate mutase (partially); saponin or Triton X-100 liberates phosphoglucose isomerase, phosphoglycerate mutase, enolase, and pyruvate kinase but not glyceraldehyde 3-phosphate dehydrogenase; freezing and thawing acts like saponin or Triton X-100 except that it fails to liberate phosphoglucose isomerase, while cell grinding with silicon carbide liberates only glyceraldehyde phosphate dehydrogenase (partially), phosphoglycerate mutase, enolase, and pyruvate kinase. The relative maximal activities of the enzymes suggest that the rate-limiting steps in glycolysis in T. brucei are the reactions catalyzed by aldolase and phosphoglycerate mutase.     

Ouabain-sensitive interaction between human red cell membrane and glycolytic enzyme complex in cytosol

Binding of 2,3-diphosphoglycerate to monophosphoglycerate mutase, of which it is an obligatory cofactor, causes changes in the resonance positions of the 31P nuclear magnetic resonance spectra of both phosphate groups. It has previously been shown that these resonances shift when other glycolytic enzymes, such as phosphoglycerate kinase, are added to form the 2,3-diphosphoglycerate . monophosphoglycerate mutase . phosphoglycerate kinase complex. In view of this association, we have examined the set of glycolytic enzymes from aldolase to pyruvate kinase and found evidence of direct communication between all of these enzymes. A multi-enzyme complex of 1--2 . 10(6) daltons has been separated from broken cell ghosts by Biogel column filtration and evidence has been presented to show that this complex exhibits aldolase, glyceraldehyde 3-phosphate dehydrogenase and phosphoglycerate kinase activity. The glycolytic multi-enzyme complex interacts with the outer face of inside-out vesicles prepared from human red cells and the interaction is suppressed by application of 10(-6) M ouabain to the inner face of these vesicles. These studies show that the conformation of the enzymes comprising the megadalton complex are responsive to the application of ouabain to the outer red cell membrane surface.     

Sensitivity of yeast glycolytic enzymes to chloroquine

Chloroquine at pH 8.0 and 10 mM concentration inhibits about 30% glucose consumption and ethanol formation in yeast cells. Out of the 11 glycolytic enzymes assayed, phosphoglycerate kinase and pyruvate decarboxylase have been found to be most sensitive to chloroquine. Next sensitive are hexokinase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase. Kinetic studies with the three kinases studied revealed competitive inhibition of chloroquine with ATP (hexokinase, phosphoglycerate kinase) or ADP (pyruvate kinase).     

The muscle-specific calmodulin-dependent protein kinase assembles with the glycolytic enzyme complex at the sarcoplasmic reticulum and modulates the activity of glyceraldehyde-3-phosphate dehydrogenase in a Ca2+/calmodulin-dependent manner

The skeletal muscle specific Ca(2)+/calmodulin-dependent protein kinase (CaMKIIbeta(M)) is localized to the sarcoplasmic reticulum (SR) by an anchoring protein, alphaKAP, but its function remains to be defined. Protein interactions of CaMKIIbeta(M) indicated that it exists in complex with enzymes involved in glycolysis at the SR membrane. The kinase was found to complex with glycogen phosphorylase, glycogen debranching enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and creatine kinase in the SR membrane. CaMKIIbeta(M) was also found to assemble with aldolase A, GAPDH, enolase, lactate dehydrogenase, creatine kinase, pyruvate kinase, and phosphorylase b kinase from the cytosolic fraction. The interacting proteins were substrates of CaMKIIbeta(M), and their phosphorylation was enhanced in a Ca(2+)- and calmodulin (CaM)-dependent manner. The CaMKIIbeta(M) could directly phosphorylate GAPDH and markedly increase ( approximately 3.4-fold) its activity in a Ca(2+)/CaM-dependent manner. These data suggest that the muscle CaMKIIbeta(M) isoform may serve to assemble the glycogen-mobilizing and glycolytic enzymes at the SR membrane and specifically modulate the activity of GAPDH in response to calcium signaling. Thus, the activation of CaMKIIbeta(M) in response to calcium signaling would serve to modulate GAPDH and thereby ATP and NADH levels at the SR membrane, which in turn will regulate calcium transport processes.     

Demonstration of a heterogeneous distribution of glycolytic enzymes and of pyruvate kinase isoenzymes types M1 and M2 in unfertilized hen eggs

The intracellular distribution of the glycolytic enzymes hexokinase, glyceraldehyde-3-phosphate dehydrogenase, lactate dehydrogenase and the pyruvate kinase isoenzymes type M1 and type M2 within unfertilized hen eggs was studied. Most of glycolytic enzyme activities were found in the yolk fraction; 8-24% of total glycolytic enzyme activities were found in the vitelline membrane fraction. However, the specific activities of these enzymes in the vitelline membrane fraction are 19-72-fold higher (U/mg protein) and 45-178-fold more concentrated (U/g wet weight) than in the yolk fraction. The study of intracellular localization of pyruvate kinase isoenzymes shows that the blastodisc, latebra and vitelline membrane contain only pyruvate kinase type M2, whereas pyruvate kinase types M1 and M2 are found in the egg yolk. The exclusive occurrence of pyruvate kinase type M2 in the blastodisc is consistent with the concept that this isoenzyme is involved in the cell proliferation. The heterogeneous distribution of the glycolytic enzymes hexokinase, glyceraldehyde-3-phosphate dehydrogenase and lactate dehydrogenase, and the heterogeneous localization of the pyruvate kinase isoenzymes types M1 and M2 indicate that glycolysis is distributed heterogeneously within the unfertilized hen egg cell.     

Glyconeogenic and glycogenic enzymes in chronically active and normal skeletal muscle

The chronically active (pseudomyotonic) gastrocnemius muscle in the C57B16J dy2J/dy2J mouse contains both elevated lactate and glycogen as well as fibers that have high amounts of glycogen and enhanced glyconeogenic activity. In the present study we analyze the activities of some key glyconeogenic enzymes to assess the causes of elevated muscle glycogen and to determine the pathway for glycogen synthesis from lactate. Glycogen synthase, malate dehydrogenase, phosphoenolpyruvate carboxykinase, and malic enzyme were all elevated in homogenates of the chronically active muscle. Activities of glycogen phosphorylase and fructose 1,6-bisphosphatase were decreased in whole muscle homogenates. Histochemistry demonstrated that the high-glycogen fibers were typically fast-twitch glycolytic fibers that had high glycogen synthase, glycogen phosphorylase, and malic enzyme activities. Malate dehydrogenase activity followed succinate dehydrogenase activity and did not correlate to high-glycogen fibers. Thus the high-glycogen fibers have an elevated enzymatic capacity for glycogen synthesis from lactate, and the pathway may involve use of the pyruvate kinase bypass enzymes.     

Interaction of immobilized phosphofructokinase with soluble muscle proteins

Selected glycolytic enzymes (including phosphoglucose isomerase, aldolase, glyceraldehyde phosphate dehydrogenase, enolase, pyruvate kinase and lactate dehydrogenase), as well as glycogen phosphorylase, creatine kinase, and adenylate kinase, bound to phosphofructokinase immobilized on an agarose gel. The affinity of phosphofructokinase to these various proteins differed, with phosphorylase exhibiting the strongest binding. Binding was reversed either by: (1) elution with high-ionic-strength buffer (0.4 M KCl); (2) the addition of a 5-10 mM concentration of ATP; or (3) high concentrations of fructose 6-phosphate (5 mM).     

Enzymes of glycolysis and the pentose phosphate pathway during development of the rabbit stomach worm Obeliscoides cuniculi

The activities of glycolytic and other enzymes of carbohydrate metabolism were measured in free-living and parasitic stages of the rabbit stomach worm Obeliscoides cuniculi. Glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, phosphoglucomutase, hexokinase, glucosephosphate isomerase, phosphofructokinase, aldolase, triosephosphate isomerase, α-glycerophosphatase, glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase, phosphoglycerate mutase, enolase, pyruvate kinase, phosphoenol pyruvate carboxykinase, lactate dehydrogenase, alcohol dehydrogenase, and glucose-6-phosphatase activities were present in worms recovered 14, 20 and 190 days postinfection.The presence of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase, and glucose-6-phosphatase indicates the possible function of a pentose phosphate pathway and a capacity for gluconeogenesis, respectively, in these worms.The ratio of pyruvate kinase (PK) to phosphoenol pyruvate carboxykinase (PEPCK) less than I in parasitic stages suggests that their most active pathway is that fixing CO₂ into phosphoenol pyruvate to produce oxaloacetate.Low levels of glucose-6-phosphate dehydrogenase, triosephosphate isomerase, PEPCK and PK were recorded in infective third-stage larvae stored at 5°C for 5 and 12 mos. The ratio of PK to PEPCK greater than 1 indicates that infective larvae preferentially utilize a different terminal pathway than the parasitic stages.     

An enzyme-coupled continuous spectrophotometric assay for glycogen synthases

The metabolic pathways leading to the synthesis of bacterial glycogen involve the action of several enzymes, among which glycogen synthase (GS) catalyzes the elongation of the α-1,4-glucan. GS from Agrobacterium tumefaciens uses preferentially ADPGlc, although UDPGlc can also be used as glycosyl donor with less efficiency. We present here a continuous spectrophotometric assay for the determination of GS activity using ADP- or UDPGlc. When ADPGlc was used as the substrate, the production of ADP is coupled to NADH oxidation via pyruvate kinase (PK) and lactate dehydrogenase (LDH). With UDPGlc as substrate, UDP was converted to ADP via adenylate kinase and subsequent coupling to PK and LDH reactions. Using this assay, we determined the kinetic parameters of GS and compared them with those obtained with the classical radiochemical method. For this purpose, we improved the expression procedure of A. tumefaciens GS using Escherichia coli BL21(DE3)-RIL cells. This assay allows the continuous monitoring of glycosyltransferase activity using ADPGlc or UDPGlc as sugar-nucleotide donors.     

Haemonchus contortus: The in vitro effects of dl-tetramisole and rafoxanide on glycolytic enzymes

Kaur R. and Sood M. L. 1982. Haemonchus contortus: the in vitro effects of dl-tetramisole and rafoxanide on glycolytic enzymes. International Journal for Parasitology 12: 585–588. Various enzymes of glycolysis (hexokinase, phosphoglucomutase, phosphoglucoisomerase, adolase, glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase, phosphoglyceromutase-enolase-pyruvate kinase and lactate dehydrogenase) have been detected in adult Haemonchus contortus. Low pyruvate kinase and lactate dehydrogenase activities suggested an alternate pathway from phosphoenolpyruvate. In vitro incubation had no significant effects on these enzymes and the worm was able to maintain normal metabolism for 12 h. Varying degrees of inhibition of glycolytic enzymes were observed with 50 μg/ml of dl-tetramisole and rafoxanide. The enzymes were inhibited to a greater extent by dl-tetramisole. These effects may block the glycolytic pathway and deprive the parasite of its ATP production.     

Haemonchus contortus: The in vitro effects of dl-tetramisole and rafoxanide on glycolytic enzymes

Kaur R. and Sood M. L. 1982. Haemonchus contortus: the in vitro effects of dl-tetramisole and rafoxanide on glycolytic enzymes. International Journal for Parasitology 12: 585–588. Various enzymes of glycolysis (hexokinase, phosphoglucomutase, phosphoglucoisomerase, adolase, glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase, phosphoglyceromutase-enolase-pyruvate kinase and lactate dehydrogenase) have been detected in adult Haemonchus contortus. Low pyruvate kinase and lactate dehydrogenase activities suggested an alternate pathway from phosphoenolpyruvate. In vitro incubation had no significant effects on these enzymes and the worm was able to maintain normal metabolism for 12 h. Varying degrees of inhibition of glycolytic enzymes were observed with 50 μg/ml of dl-tetramisole and rafoxanide. The enzymes were inhibited to a greater extent by dl-tetramisole. These effects may block the glycolytic pathway and deprive the parasite of its ATP production.