SEARCH · PubMed Health

Results for “HEXOKINASE”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 505 records · Page 28Linked to original sources

PubMed1982

[Interaction of isozymes I and II of hexokinase with a microsomal fraction of skeletal muscle of rats].

The conditions of reversible transition of hexokinase from the hyaloplasm to the rat skeletal muscle microsomal membranes were studied. The maximal absorption of the enzyme by the membranes was observed in the presence of Mg2+ and glucose at weakly alkaline values of pH. Adenyl nucleotides and glucose 6-phosphate taken at physiological concentrations at neutral and weakly acid values of pH make the effect of Mg2+ reversible. It was shown that the both hexokinase isozymes I and II localized in muscle hyaloplasm are capable of reversible transition from the free state to the bound one. All the effects observed during hexokinase interaction with the rough microsomal fraction occurred in the cases of a purified muscle sarcoplasmic reticulum fraction. The conditions for the enzyme interaction with the microsomal and mitochondrial membrane fractions were found to be practically identical. However, when the enzyme was bound to the microsomal membrane, some reversible changes (e. g. increase of V) took place; the Km value for glucose remained thereby unchanged. The data obtained are discussed in terms of an adsorption mechanism of hexokinase activity control in the cell.

Animals↗

PubMed1981

Influence of thiopental on intracellular distribution of hexokinase activity in various tumor cells.

In earlier work it has been shown that mitochondrially bound brain hexokinase is solubilized by anesthetics. This effect was reevaluated using cultured cells. For the present experiments Ehrlich ascites, Harding-Passey melanoma, C-1300-neuroblastoma and C-6-glioma cells were used. The great portion of hexokinase activity bound to the mitochondria of these cells was similar to that in rat brain. After incubation with thiopental the soluble hexokinase activity was increased in all cells studied. Using neuroblastoma and glioma cells the thiopental effect was demonstrated to be dose-dependent. Thus, cultured tumor cells seem to be useful for studying the relationship of the intracellular distribution of hexokinase activity, energy metabolism and the effect of anesthetics.

Cell Division↗

PubMed1980

Rabbit red blood cell hexokinase. Purification and properties.

Rabbit red blood cell hexokinase (EC 2.7.1.1.) has been purified 300,000-fold by a combination of ion exchange chromatography, affinity chromatography, and preparative polyacrylamide gel electrophoresis. The hexokinase activity has been isolated in 35% yield as a protein that is homogeneous by polyacrylamide and sodium dodecyl sulfate gel electrophoresis. The highest specific activity obtained was 145 units/mg of proteins. The native protein has a molecular weight of 110,000 by gel filtration on Ultrogel AcA 44 and 112,000 by sedimentation velocity on sucrose density gradients. Sodium dodecyl sulfate-polyacrylamide gels gave a molecular weight of 110,000 indicating that hexokinase is a monomer. The enzyme had a pI of 6.20 to 6.30 pH units by isoelectric focusing. The enzyme was specific for Mg . ATP and Mg . ITP as the nucleotide substrates. Several hexokinase with different affinities.

Adenosine Triphosphate↗

PubMed1994

Glycolysis and growth rate in normal and in hexokinase-transfected NIH-3T3 cells.

The glycolytic enzyme hexokinase plays a key role in regulating cell energy metabolism. Its activity has been associated with cell growth rate and, notably, with neoplastic transformation. NIH-3T3 cells were transfected with a tumor hexokinase cDNA. The transfected cells showed increased hexokinase amount and activity, mainly located in the particulate cellular fraction, increased glycolytic rate evaluated as lactate production, and, finally, enhanced growth rate. These data may suggest that high hexokinase activity might be not merely the consequence of peculiar metabolic demands by actively replicating normal or cancer cells, but also a modification able per se to drive, at least partially, a more intense mitotic activity.

3T3 Cells↗

PubMed1994

[Solubilization of mitochondria-bound rat skeletal muscle hexokinase isoenzyme II by glucose-6-phosphate].

Solubilization of bound to outer mitochondrial membrane hexokinase isoenzyme II by glucose-6-phosphate (G-6-P) has been studied. Dissociation of the enzyme-membrane complex was analyzed in both active (in the presence of reaction substrates) and inactive states. The magnitude of the complex dissociation constants was determined under G-6-P action in the presence and absence Mg.ATP. Besides, some kinetic parameters of hexokinase isoenzyme II in its free and bound forms were estimated. It is suggested that the mechanism of dissociation of the hexokinase-membrane complex by G-6-P is coupled with the kinetic mechanism of the reaction catalyzed by hexokinase isoenzyme II.

Animals↗

PubMed1996

Glucose catabolism in cancer cells: amplification of the gene encoding type II hexokinase.

Hexokinase type II is highly overexpressed in many cancer cells, where it plays a pivotal role in the high glycolytic phenotype. Here we demonstrate by Southern blot analysis and fluorescence in situ hybridization (FISH) that in the rapidly growing rat AS-30D hepatoma cell line, enhanced hexokinase activity is associated with at least a 5-fold amplification of the type II gene relative to normal hepatocytes. This amplification is located chromosomally, extends to the whole gene, and most likely occurs at the site of the resident gene. No rearrangement of the gene could be detected. Therefore, overexpression of hexokinase type II in AS-30D hepatoma cells may be based, at least in part, on a stable gene amplification. This is the first report describing the amplification of a hexokinase gene in a tumor cell line expressing the high glycolytic phenotype.

Animals↗

PubMed1995

[Isoenzymatic characterization of the hexokinase in the development of Plebeia droryana (Hymenoptera, Apidae) and its relation with flight activity].

Hexokinase enzyme was studied by means of starch-agarose gel electrophoresis in Plebeia droryana. Three anodal bands with enzyme activity were observed during the development: hexokinase-1 (HK-1), the intensity of which increases from larvae to adult, probably related with energy supply to thoracic flight muscles and, therefore, with flight activity; hexokinase-2 (HK-2), that reaches maximum intensity in clear brown eyed pupae and hexokinase-3 (HK-3), the intensity of which reaches maximum peak in winged pupae and is not observed in the adult phase. This isozyme should have important functions in the bee metamorphosis.

Animals↗

PubMed1983

Subcellular localization of hexose kinases in pea stems: mitochondrial hexokinase.

The subcellular localization of hexose phosphorylating activity in extracts of pea stems has been studied by differential centrifugation and sucrose density gradient centrifugation. The hexokinase (EC 2.7.1.1) was associated with the mitochondria, whereas fructokinase (EC 2.7.1.4) was in the cytosolic fraction. Some properties of the mitochondrial hexokinase were studied. The enzyme had a high affinity for glucose (K(m) 76 micromolar) and mannose (K(m) 71 micromolar) and a relatively low affinity for fructose (K(m) 15.7 millimolar). The K(m) for MgATP was 180 micromolar. The addition of salts stimulated the activity of the hexokinase. Al(3+) was a strong inhibitor at pH 7 but not at the optimum pH (8.2). The enzyme was not readily solubilized but, in experiments with intact mitochondria, was susceptible to proteolysis. A location on the outer mitochondrial membrane is suggested for the hexokinase of pea stems.

Journal Article↗

PubMed2001

Regulation of hexokinase in a freeze avoiding insect: role in the winter production of glycerol.

Hexokinase from larvae of the freeze-avoiding goldenrod gall moth, Epiblema scudderiana, was purified 20-fold using chromatography on DE52 Sephadex, phosphocellulose, and blue dextran. Final specific activity was 75.8 U/mg and SDS-PAGE gave a molecular weight of 94,000 for the monomer. Arrhenius plot showed a break at 16 degrees or 12 degrees C in the absence vs. presence of 10% v/v glycerol, indicating a conformational change in the enzyme at lower temperatures but suggesting a stabilizing effect of glycerol. Comparison of hexokinase kinetic properties at 22 degrees and 4 degrees C showed higher affinity for both glucose and ATP (Km values were 45-50% lower), as well as for the cofactor Mg(2+), at the lower temperature. Furthermore, product inhibition by glucose-6-phosphate and ADP was reduced at 4 degrees C. Glucose levels rise in E. scudderiana as an apparent by-product of high rates of glycogenolysis during glycerol synthesis. The temperature-dependent properties of hexokinase would facilitate the recycling of this glucose back into the pathway of glycerol synthesis and could help to achieve the near stoichiometric conversion of glycogen to glycerol that is seen during cold hardening. Arch.

Animals↗

PubMed1994

Red blood cell phagocytosis following hexokinase inactivation.

Hexokinase inactivating antibodies were loaded into human erythrocytes using an encapsulation procedure based on hypotonic haemolysis, isotonic resealing and reannealing. Red blood cells loaded with anti-hexokinase IgG showed 20 percent residual hexokinase activity and reduced glycolytic activity. Incubation of these cells in the presence of an oxidizing agent such as terbutyl hydroperoxide (TBH) and then in autologous plasma, promoted opsonization by autologous IgG and complement deposition, but not haemolysis. Furthermore, the antihexokinase IgG loaded cells treated with TBH were actively recognized and phagocytosed by macrophages. Thus, metabolic impairment of human erythrocytes promotes autologous IgG binding, C3 deposition and phagocytosis, a mechanism already reported for the removal of senescent erythrocytes.

Complement System Proteins↗

PubMed1993

The structure of the MnIIADP-nitrate-lyxose complex at the active site of hexokinase.

The coordination scheme of Mn2+ in the hexokinase-MnIIADP-nitrate-lyxose complex has been determined by electron paramagnetic resonance (EPR) spectroscopy with 17O-enriched ligands. Nitrate binds to the active site of hexokinase when MnIIADP and a sugar substrate or analogue are present. The binding of nitrate enhances inhibition by glucose when ADP is present and narrows the EPR signals of the enzyme-bound MnIIADP complex in the presence of sugar substrates or analogues. Experiments using regiospecifically 17O-enriched ADP, 17O-enriched nitrate, and 17O-enriched water establish the coordination scheme of Mn2+. The EPR experiments show that ADP is a beta-monodentate ligand and that nitrate binds directly to Mn2+. Four water molecules complete the coordination sphere of the enzyme-bound Mn2+. The dissociation constant (Kd approximately 8 mM) of nitrate for the complex with enzyme, MnIIADP, and lyxose was obtained from titration experiments. These results suggest that nitrate-stabilized, dead-end complexes of hexokinase may be useful in stabilizing the closed conformation of this "hinge-bending" enzyme for crystallographic experiments.

Adenosine Diphosphate↗

PubMedDate not supplied

Downeast anemia (dea), a new mouse model of severe nonspherocytic hemolytic anemia caused by hexokinase (HK(1)) deficiency.

A new spontaneous mutation in the A/J inbred mouse strain, downeast anemia (dea), causes severe hemolytic anemia with extensive tissue iron deposition and marked reticulocytosis. The anemia is present at birth and persists throughout life. The defect is inherited as an autosomal recessive and is transferable through bone marrow stem cells. The red cell morphology is consistent with a nonspherocytic hemolytic anemia, suggestive of a red cell enzymopathy. In linkage analysis, dea is nonrecombinant with the hexokinase-1 gene (Hk1) on mouse Chromosome 10. Expression of Hk1 is markedly decreased in dea erythroid tissues, and the transcript produced is larger than normal. Hexokinase enzyme activity is significantly decreased in dea tissues, including red cells, spleen, and kidney. Southern blot analyses revealed approximately 5.5 kb of additional sequence in the 5' portion of the dea Hk1 gene, which was identified by direct sequencing as an early transposon (ETn) insertion in intron 4. ETn insertions disrupt genes in several mouse models by a variety of mechanisms, including aberrant splicing of ETn sequences into the mRNA. We conclude that the primary gene defect in the dea mutation is in Hk1 and that dea is a model of generalized hexokinase deficiency, the first such model identified to date.

Anemia, Hemolytic↗

PubMed1996

High-level expression and purification of a human "mini"-hexokinase.

Human hexokinase type I is a 100-kDa enzyme with the catalytic site located in the C-terminal domain. We had previously expressed this domain in Escherichia coli, however only a small amount of the recombinant enzyme was catalytically active. To overcome this problem we have now expressed the "mini"-hexokinase using the pET expression system. An average of 1000 U of enzyme per liter of culture was obtained. The recombinant enzyme was purified to homogeneity by a combination of ion-exchange chromatography, affinity chromatography, and dye-ligand chromatography. The enzyme was unstable under ultrafiltration; thus, a multicolumn purification procedure was developed in order to avoid the ultrafiltration steps. The recombinant "mini"-hexokinase was found to have the same kinetic properties as the entire enzyme. Using the method described, the enzyme can be obtained in sufficient quantities for biophysical and biochemical investigations.

Blotting, Western↗

PubMed1977

An inverse relation between mitochondrial hexokinase content and phosphoglucomutase activity of rat tissues.

The hexokinase: fumarase ratios of mitochondria isolated from ten tissues of the rat were determined, and compared with the tissue content of phosphoglucomutase and phosphorylase, taken as representatives of enzymes concerned with glycogen metabolism. A generally inverse relationship was found between the mitochondrial hexokinase: fumarase ratio and phosphoglucomutase levels. The cytochrome: fumarase ratios were relatively invariant in these same mitochondria. The results are interpreted as indicating a specialization of mitochondria, with increased amounts to hexokinase being associated with the mitochondria in tissues exhibiting less dependence on glycogen metabolism, as judged from phosphoglucomutase levels.

Animals↗

PubMed1990

Differential energetic metabolism during Trypanosoma cruzi differentiation. II. Hexokinase, phosphofructokinase and pyruvate kinase.

The activities of hexokinase (ATP:hexose-6-phosphate transferase, E.C. 2.7.1.1), phosphofructokinase (ATP:fructose-6-phosphate 1-phosphotransferase, E.C.2.7.1.11) and pyruvate kinase (ATP:pyruvate transferase, E.C. 2.7.1.40), and their kinetic behaviour in two morphological forms of Trypanosoma cruzi (epimastigotes and metacyclic trypomastigotes) have been studied. The kinetic responses of the three enzymes to their respective substrates were normalized to hyperbolic forms on a velocity versus substrate concentration plots. Hexokinase and phosphofructokinase showed a higher activity in epimastigotes than in metacyclics, whereas pyruvate kinase had similar activity in both forms of the parasite. The specific activity of hexokinase from epimastigotes was 102.00 mUnits/mg of protein and the apparent Km value for glucose was 35.4 microM. Metacyclic forms showed a specific activity of 55.25 mUnits/mg and a Km value of 46.3 microM. The kinetic parameters (specific activity and Km for fructose 6-phosphate) of phosphofructokinase for epimastigotes were 42.60 mUnits/mg and 0.31 mM and for metacyclics 13.97 mUnits/mg and 0.16 mM, respectively. On the contrary, pyruvate kinase in both forms of T. cruzi did not show significant differences in its kinetic parameters. The specific activity in epimastigotes was 37.00 mUnits/mg and the Km for phosphoenolpyruvate was 0.47 mM, whereas in metacyclics these values were 42.94 mUnits/mg and 0.46 mM, respectively. The results presented in this work, clearly demonstrate a quantitative change in the glycolytic pathway of both culture forms of T. cruzi.

Animals↗

PubMed1992

Cleavage of hexokinase II to two domains by trypsin without significant change in catalytic activity.

Hexokinase II prepared from Ehrlich-Lettre hyperdiploid tumor cells (ELD cells) was subjected to a limited digestion by trypsin. After 60 min digestion, hexokinase II (100 kDa) was completely cleaved to two fragments with the molecular weight of about 60 kDa and 40 kDa as manifested in SDS-PAGE. It was noteworthy that the enzyme activity was observed even at the time when the native enzyme molecule was no more detectable. These fragments were separated by SDS-PAGE irrespective of the presence of a reducing agent, but neither by native PAGE nor by cellulose acetate membrane electrophoresis under the nondenaturing conditions. Neither kinetic parameters such as Km values for ATP and glucose nor an ability of binding to mitochondria were changed significantly by the tryptic digestion. These results indicate that an essential conformation of hexokinase II can be restored by the self-association of two fragments produced as a result of the cleavage by trypsin at the middle of the molecule. Affinity labeling with 2',3'-dialdehyde ATP followed by the trypsin digestion showed that ATP binding site resided in the 40 kDa fragment. Furthermore, the mode of the response in the incorporation of this ATP analog to hexose phosphate, moreover, was similar to that in the catalytic activity.

Animals↗

PubMed1995

Polymorphisms of the human hexokinase II gene: lack of association with NIDDM and insulin resistance.

Skeletal muscle and adipose tissue hexokinase II is a promising candidate gene for non-insulin-dependent diabetes mellitus (NIDDM) and insulin resistance. Therefore, we investigated the association of alleles at four polymorphic loci in this gene with NIDDM and insulin resistance in 110 Finnish diabetic patients with NIDDM and in 97 Finnish control subjects with normal glucose tolerance and a negative family history of diabetes. The four polymorphic nucleotide substitutions (silent) in the coding region of the hexokinase II gene were: GAC 251 GAT (exon 7), AAC 692 AAT and CCG 736 CCC (exon 15), and CTG 766 CTA (exon 16). Allele frequencies of each of these polymorphisms did not differ between patients with NIDDM and control subjects. In addition, subjects who were homozygous for the less frequent allele of each of the four polymorphisms had a similar degree of insulin resistance, as determined by the euglycaemic clamp technique, as did the subjects who were homozygous for the common allele in both control subjects and in patients with NIDDM. In conclusion, polymorphisms in the hexokinase II gene are not associated with the risk of NIDDM or insulin resistance in the Finnish population.

Adipose Tissue↗

PubMed1990

Studies of hexokinase activity in human and mouse oocyte.

Microdetermination methods were used to determine the activities of hexokinase in human and mouse oocytes, human spermatozoa, and other somatic cells. Human oocytes with intact germinal vesicle obtained from the growing follicle were freeze dried and weighed (mean +/- SD = 243 +/- 34 ng dry weight). Hexokinase activity in single oocytes was 17.9 +/- 3.6 pmol of reduced nicotinamide-adenine dinucleotide phosphate formed/oocyte/hr at 20 degrees C. Specific activity was estimated to be 104 +/- 30 nmol/mg protein/hr, which was remarkably lower than that in human spermatozoa (3570 +/- 550 nmol/mg protein/hr) and other somatic cells such as endometrium, brain, liver, and kidney. Mouse follicular oocytes and early embryos before the two-cell stage also had low hexokinase activities, but morulae and blastocysts had increasingly high activities; the former cannot use glucose as an energy source, whereas the latter can. These results suggest that human immature oocytes, as well as mouse oocytes, depend on pyruvate instead of glucose as a major energy source.

Adult↗