Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.1.1 (hexokinase)
5,274 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Glutathione S-transferase (GST) purified from Schistosoma mansoni or human placenta was inhibited by the antischistosomal drug oltipraz (OPZ) in a time- and concentration-dependent manner. Inhibition of placenta GST was complete at a low concentration of drug, whereas that of parasite GST was incomplete and relatively high amounts of OPZ were needed to reach 50% inhibition. Complete reactivation of GST from placenta was achieved with dithiothreitol (DTT) and other sulfhydryl-containing compounds, while the inactivation of parasite GST was irreversible. The oxy-derivative of OPZ (RP 36,642), in which the thione sulfur is replaced with oxygen, did not inhibit GST activity. There were no differences between OPZ and RP 36,642 in their patterns of binding to the hydrophobic non-substrate site of GST. GST from the placenta incorporated much higher levels of [14C]N-ethylmaleimide compared to schistosome GST. The incorporation of [14C]N-ethylmaleimide by GST was inhibited by OPZ but not by RP 36,642. Yeast and S. mansoni hexokinases were similarly inhibited by OPZ but not by RP 36,642. Both hexokinase preparations recovered their activity following incubation with DTT. These data suggest that the inactivation of these enzymes by OPZ is a result of its interaction with their SH groups. Thus, the antischistosomal activity of OPZ may be accounted for by its interaction with the SH groups of macromolecules in general.
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PMID:Mechanisms of inactivation of Schistosoma mansoni and mammalian glutathione S-transferase activity by the antischistosomal drug oltipraz. 156 85

5-Thioglucose (5-TG) had a marked effect on the energy metabolism of Schistosoma mansoni in vitro: the conversion of external glucose into lactate by intact worms was severely inhibited. This inhibition of glycolysis was instantaneous, independent of the oxygen concentration and competitive with respect to glucose. Degradation of 0.5 mM external (14C-labelled) glucose was inhibited for 80% in the presence of 20 mM 5-TG. On the other hand the degradation of endogeneous glycogen to lactate was uninhibited. This shows that the inhibition of glucose breakdown occurred at the entrance of glucose into the cell and/or at the hexokinase reaction. It was demonstrated that 5-TG inhibited both the uptake of glucose and the activity of hexokinase. However, it was concluded that in the intact worm 5-TG blocked glycolysis by its competitive inhibition of hexokinase. In intact S. mansoni worms hexokinase is probably the rate-limiting enzyme of glycolysis. Krebs-cycle activity and lactate production do not occur at a fixed ratio: at lower rates of pyruvate formation Krebs-cycle activity was favoured.
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PMID:The effect of 5-thioglucose on the energy metabolism of Schistosoma mansoni in vitro. 403 43

The enzyme phosphoglucose isomerase (PGI), phosphoglucomutase (PGM), hexokinase (HK), adenylate kinase (AK), fructokinase (FK), mannose-6-phosphate isomerase (MPI), glucose-6-phosphate dehydrogenase (G-6-PDH) and malate dehydrogenase (MDH) were chosen to study the variation between isolates, cercariae and adults, individuals, and sexes of Schistosoma mansoni and S. rodhaini, using horizontal polyacrylamide gel electrophoresis. The method described allows combinations of six of the eight enzymes to be scored in the homogenate from one adult worm. In adult S. mansoni one phenotype of the eight enzymes was observed in all isolates. In addition, the enzyme PGI showed polymorphism in the isolates from Tala, Kenya and Uganda. PGM in the isolates from Tala, Kenya and South Africa showed polymorphism. The cercarial phenotype differs from the adult phenotype in G-6-PDH, where the cercarial enzyme mobility is slower than that in the adult worm. The low amount of intrastrain variation observed in this species is explained by the limited amount of material used to establish the laboratory stocks, whereas the genetic similarity between geographically widely separated stocks does suggest that only limited geographical variation is likely to occur in S. mansoni. It is suggested that the gene controlling the PGI polymorphism is located on the sex chromosomes of S. mansoni. Mobility differences were observed between S. mansoni and S. rodhaini in the enzyme PGI and PGM, and these characteristics might be useful for a quick identification of schistosome cercariae emerging from Biomphalaria sp. in Africa.U
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PMID:Isoenzyme studies on cercariae from monoinfections and adult worms of Schistosoma mansoni (10 isolates) and S. rodhaini (one isolate) by horizontal polyacrylamide gel electrophoresis and staining of eight enzymes. 621

DNA encoding a Schistosoma mansoni hexokinase (SHEX) was amplified from cDNA by the polymerase chain reaction using opposing oligonucleotide primers designed to hybridize with two short segments of hexokinase coding sequences that are well-conserved through evolution. The resulting DNA fragment was then used as a probe to identify a full-length hexokinase cDNA clone. SHEX cDNA encodes a 50-kDa protein that is approximately 46% homologous to rat hexokinase, 40% to rat glucokinase, and 34% to yeast hexokinase A. SHEX coding DNA was expressed within Escherichia coli cells and the 50-kDa recombinant product (rSHEX) was partially purified. Mice repeatedly immunized with rSHEX produced antibodies which recognize rSHEX but this offered no significant protection against subsequent cercarial challenge. On Western blots, rSHEX is weakly recognized by antisera against rat brain hexokinase but not by sera from three strains of mice experimentally infected with S. mansoni parasites or from numerous human schistosomiasis patients. Thus, unlike other reported S. mansoni glycolytic enzymes, hexokinase appears to be poorly immunogenic during schistosome infection and of limited potential as a vaccine candidate.
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PMID:Schistosoma mansoni hexokinase: cDNA cloning and immunogenicity studies. 782 9

Hexokinase has been purified from adult Schistosoma mansoni worms and the activity shown to be associated with a single protein species having an M(r) about 50,000. This protein is recognized on Western blots probed with antisera against rat Type I hexokinase or against a recombinant S. mansoni hexokinase that had been expressed in Escherichia coli using a previously cloned cDNA. An 18-residue N-terminal sequence determined for the purified S. mansoni hexokinase is identical to that deduced from the nucleotide sequence of the cDNA, consistent with the view that the cloned cDNA encodes the hexokinase characterized in the present study. The S. mansoni enzyme has a relatively low Km (approximately 60 microM) for glucose and is sensitive to inhibition (competitive versus ATP, Ki approximately 50 microM) by its product, glucose 6-phosphate (Glc-6-P). With these kinetic properties and 50 kDa molecular mass, S. mansoni hexokinase resembles the ancestral hexokinase predicted to have given rise, by gene duplication and fusion, to the present day 100-kDa Glc-6-P-sensitive mammalian hexokinases. The schistosomal hexokinase represents the first 50-kDa Glc-6-P-sensitive hexokinase whose sequence has been obtained. The schistosomal hexokinase does not bind to mitochondria, consistent with its lack of a hydrophobic segment at the N terminus which is required for binding of the mammalian Type I and II isoenzymes to mitochondria. The marked Crabtree effect exhibited by S. mansoni cercariae may be at least partly attributed to the expression of rather high levels of a hexokinase having a high affinity for glucose but only a moderate sensitivity to product inhibition by Glc-6-P.
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PMID:The 50-kDa glucose 6-phosphate-sensitive hexokinase of Schistosoma mansoni. 792 49

Schistosomes switch rapidly from the use of stored glycogen to a reliance on host glucose during the transformation from free-living cercariae to parasitic schistosomula. We have cloned a set of cDNAs encoding proteins involved in glucose metabolism to allow us to examine the expression of these genes during this transformation. We first obtained and characterized Schistosoma mansoni cDNA clones encoding the tricarboxylic acid cycle enzyme, mitochondrial malate dehydrogenase (SMDH) and the mitochondrial encoded electron transport protein, cytochrome oxidase subunit 1 (SCOX1). Northern blots were then prepared using mRNA isolated from whole cercariae, cercarial tails, schistosomula, adult males and adult females. The Northern blots were successively hybridized with a variety of probes including those for SMDH, SCOX, the glycolytic enzymes, hexokinase, triosephosphate isomerase and glyceraldehyde-3-phosphate dehydrogenase and several control probes. Probes were additionally hybridized to mRNA dot blots and the signals were quantified using storage phosphor technology. These studies reveal that transcripts encoding these metabolic enzymes are localized at much higher levels in cercarial tails than in whole cercariae or transformed schistosomula, and support the notion of a dominant aerobic metabolism in tails. Male and female adult worms express each of the mRNAs at roughly equal levels. Adults express the metabolic mRNAs, including those involved in oxidative glucose metabolism, at relatively high levels suggesting that adult schistosomes retain a significant capacity to produce energy through aerobic metabolism.
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PMID:Expression of Schistosoma mansoni genes involved in anaerobic and oxidative glucose metabolism during the cercaria to adult transformation. 839 6

An ATP-diphosphohydrolase (EC 3.6.1.5) was identified in the tegumental fraction isolated from Schistosoma mansoni worms. Both ATP and ADP were hydrolyzed to AMP at similar rates by the enzyme. Other nucleotides were also degraded by the tegument enzyme, revealing a broad substrate specificity. Electrophoretic separation of tegumental proteins under non-denaturing conditions followed by addition of ATP or ADP as substrate revealed a single band of activity with similar mobility. In addition, similar heat-inactivation profiles were obtained for ATPase or ADPase activities, indicating that a single enzyme is responsible for degrading both nucleotides. The enzyme was not inhibited by vanadate, levamisole, tetramisole, ouabain or sodium azide. The ADPase activity was not affected by adenosine (5')-pentaphospho-(5')-adenosine (Ap5A) or by an excess of glucose and hexokinase used as an ATP-trapping system, thus excluding the presence of any significant adenylate kinase activity. The ATP-diphosphohydrolase displayed micromolar affinities for both Mg2+ and Ca2+, and the calcium-activated enzyme was inhibited by millimolar Mg2+. In intact live worms a calcium phosphate precipitate was formed on the outer tegumental surface upon incubation of the worms with either ATP or ADP, indicating the ectolocalization of this enzyme. In addition, ultrastructural histochemical localization of the enzyme was obtained. A distinct deposition of lead phosphate granules on the outer surface of the tegument was observed by electron microscopy, in the presence of either ATP or ADP as substrate. It is suggested that the ATP-diphosphohydrolase could regulate the concentration of purine nucleotides around the parasites and hence enable them to escape the host hemostasis by preventing ADP-induced platelet activation.
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PMID:Characterization and localization of an ATP-diphosphohydrolase on the external surface of the tegument of Schistosoma mansoni. 847 45

The hexokinase (ATP;D-hexose 6-phosphotransferase, EC 2.7.1.1) of Schistosoma mansoni has been expressed in Escherichia coli as a fusion protein including an N-terminal polyhistidine tag and enterokinase cleavage site. The enzyme was purified by metal chelate chromatography to > 95% homogeneity, based on analysis by SDS-gel electrophoresis and isoelectric focusing. The absorbance at 280 nm was 0.54 for a 1 mg/ml solution (molar extinction coefficient 2.7 x 10(4) cm2 mol). The pI of the S. mansoni hexokinase was 6.0-6.2, slightly more acidic than the rat Type I isozyme (pI 6.35). The S. mansoni enzyme migrated as a single band of activity during nondenaturing cellulose acetate electrophoresis; the mobility was slightly greater than the rat Type I isozyme, consistent with the estimated pI. The Km values for substrates glucose and ATP were 128 +/- 10 and 927 +/- 41 microM, respectively. In accord with a previous report, the S. mansoni hexokinase exhibited moderate sensitivity to inhibition (competitive vs ATP) by the product, glucose 6-phosphate, with a Ki approximately 150 microM; the product analog, 1,5-anhydroglucitol 6-phosphate, was somewhat less effective as an inhibitor, with Ki approximately 500 microM. These kinetic properties were not altered by removal of the N-terminal fusion partner by enterokinase treatment. Immunological crossreactivity between the rat Type I isozyme and the S. mansoni hexokinase was demonstrated by immunoblotting, but this was markedly dependent on the preparation of antiserum used. The activity of the enzyme is apparently highly dependent on maintenance of free sulfhydryl groups. Activity was maintained during storage in the presence of monothioglycerol; activity lost during storage in the absence of monothioglycerol could be partially restored by treatment with this reagent.
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PMID:Purification and characterization of the hexokinase from Schistosoma mansoni, expressed in Escherichia coli. 893

Studies on human and murine isolates of Schistosoma mansoni, from Richard-Toll, Senegal, were carried out by isoelectric focusing in polyacrylamide gels. Seven enzyme systems; lactate dehydrogenase (LDH), malate dehydrogenase (MDH), glucose-6-phosphate dehydrogenase (G6PD), acid phosphatase (AcP), hexokinase (HK), glucose phosphate isomerase (GPI), and phosphoglucomutase (PGM), were used to compare the two isolates. All systems tested, apart from LDH, were found to be polymorphic for both isolates. Interestingly, one phenotype is more frequent than the remainder. The results show that there is no significant genetic variation between the S. mansoni isolates from man and the rodents, Arvicanthis niloticus and Mastomys huberti.
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PMID:Comparison of human and murine isolates of Schistosoma mansoni from Richard-Toll, Senegal, by isoelectric focusing. 919 7

We have determined the structures of the glucose-6-phosphate (G6P)-inhibitable 100,000 Mr Type I hexokinase from rat and the G6P-sensitive 50,000 Mr hexokinase from Schistosoma mansoni at a resolution of 2.8 and 2.6 A respectively. The structures define the glucose and G6P binding sites in these enzymes, suggest the mechanisms of intradomain G6P inhibition and activity loss in the Type I hexokinase N-terminal half, and reveal the structure of the membrane targeting motif that integrates the Type I hexokinase into the outer mitochondrial membrane.
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PMID:The structure of mammalian hexokinase-1. 966 68


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