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)

Yeast hexokinase A(ATP:D-hexose 6-phosphotransferase) is inactivated when incubated in the presence of xylose and ATPMg, or in the presence of D-lyxose in a reaction medium in which ATPMg is being continuously regenerated (phosphoenolpyruvate and pyruvate kinase). The inactivation is due to the phorphorylation of the protein. A linear relationship was observed between the inactivation and the incorporation of 32P from [gamma-32P] ATP. All hexokinase and ATPase activity of the enzyme is lost when one phosphoryl group is incorporated per enzyme subunit (molecular weight 51,000). The phosphoryl group is covalently bound by a ester linkage with a serine residue of the protein.
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PMID:Studies on the active site of yeast hexokinase. Specific phosphorylation of a serine residue induced by D-xylose and ATPMg. 0 82

A protein phosphokinase (EC 2.7.1.1.37) was isolated from baker's yeast (Saccharomyces cerevisiae) after a 17,000-fold purification; the purified enzyme is homogeneous according to the criteria of gel electrophoresis and ultracentrifuge analysis. The enzyme has a high isoelectric point of ca. 9 and appears to exist as a monomer with a molecular weight of 42,000 plus or minus 1500. It is neither stimulated by cyclic 3',5'-AMP, -GMP, -CMP or -ump nor inhibited by the regulatory subunit of rabbit muscle protein kinase (Reimann, E. M., Walsh, D. A., and Krebs, E. G. (1971), J. Biol. Chem. 246, 1986). In the presence of divalent metal ions, preferably Mg-2+ or Mn-2+, the enzyme readily transfers the terminal phosphate group of ATP to phosvitin, alphaS1B- and beta a-casein and an NH2-terminal tryptic peptide derived from beta a-casein, but not to protamine, lysine, or arginine-rich histones or to yeast enzymes such as phosphorylase, phosphofructokinase, or pyruvate carboxylase; serine and polyserine were also inactive as phosphate acceptors. Km values of 0.17 mM for beta a-casein and 0.2 mMfor ATP were determined at 10 mM Mg-2+. The urified yeast protein kinase also catalyzes the reverse reaction, namely, the transfer of phosphate from fully phosphorylated beta a-casein or its NH2-terminal peptide to ADP resulting in the formation of ATP. AMP, GDP, UDP, and CDP did not serve as phosphate acceptors in this reaction. As observed by Rabinowitz and Lipmann (Rabinowitz, M., and Lipmann, F. (1960), J. Biol. Chem. 235, 1043) both reactions have different pHoptima with values of 7.5 for the forward reaction (phosphorylation of the proteins) and ca 5.2 for the formation of ATP; both are differently affected by salts. Phosphorylation of beta a-casein with [gamma-32-P]ATP followed by digestion of the labeled protein with trypsin indicated that all the radioactivity was exclusively introduced in an NH2-terminal peptide possessing the unique sequence: Glu-Ser(P)-Leu-Ser(P)-Ser(P)-Ser(P)-Glu-Glu...(Ribadeau-Dumas, B., Brignon, G., Grosclaude, F., and Mercier, J.-C. (1971), eur J. Biochem. 20, 264). By subjecting beta a-casein and its NH2-terminal peptide to the combined action of almond acid phosphatease and purified yeast protein kinase, it was determined that the phosphorylation and dephosphorylation reactions proceed randomly, i.e., all seryl phosphate residues are equally susceptible and that the rate of phosphorylation decreases drastically as the number of bound phosphate groups in the substrate diminishes.
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PMID:Purification and properties of a yeast protein kinase. 23 75

As a common characteristic of tumor cells, as well as of normal proliferating cells in the G1-phase of cell cycle, one finds constitutive high levels of all the glycolytic metabolites arising between glucose 6-phosphate and phosphoenolpyruvate. Thus, it is that the phosphometabolites fructose 1,6-bisphosphate, ribose 5-P, P-ribose-PP, NAD, GTP, CTO, UTP, UDP-glucose, glycerol 3-P, glycerol phosphocholine and glycerol phosphoethanolamine are useful in the 31P-nuclear magnetic resonance (NMR) detection of solid tumors in animals and man. This expansion of phosphometabolites is achieved during tumor formation as a result of reductions in levels of enzymes degrading phosphometabolites, owing to the decline in the glycerol 3-P hydrogen shuttle, and as a consequence of alterations in the glycolytic isoenzyme equipment. Tumor cells typically express a particular isoenzyme of pyruvate kinase called type M2 (K) at high levels. This isoenzyme is subject to a complex regulation by amino acids, by fructose 1,6-bisphosphate, and by hormonal- and oncogene-dependent phosphorylation. Pyruvate kinase type M2 is a substrate for the oncogene encoded PP60v-src-tyrosine kinase. A drastic decrease in the affinity for its substrate phosphoenolpyruvate found after transformation by the src-oncogene can be explained as a consequence of the phosphorylation of pyruvate kinase in serine and tyrosine. These phosphorylations induce the breakdown of tetrameric pyruvate kinase to the trimeric and dimeric forms. Unlike the tetrameric form, the dimeric form as a low affinity for phosphoenolpyruvate. Partial inactivation of pyruvate kinase and enolase on the one hand, and a hyperactivation of hexokinase and phosphofructokinase on the other hand, lead to an expansion of all metabolites. Only when these metabolites attain high levels, thereby assuring a sufficient supply of metabolites for RNA, DNA, lipid, and complex carbohydrate synthesis, can cell proliferation proceed. This accumulation of metabolites in the G1-phase cells has been termed a "metabolic budget system" because it senses not only the actual nutrient levels, but also the supply over a period of time. Monoclonal antibodies specific for the dimeric form of pyruvate kinase type M2 can be used for the immunohistological detection of tumor cells. The amount of the dimeric form in tumor cells closely correlates with the degree of malignancy and can be used for a nonspecific detection of tumors based on assays performed with patient's plasma.
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PMID:Double role for pyruvate kinase type M2 in the expansion of phosphometabolite pools found in tumor cells. 153 31

Mammalian hexokinase type I is a 100 kDa enzyme that has been considered to be evolved from an ancestral 50 kDa yeast-type hexokinase, insensitive to product inhibition, by gene duplication and fusion. According to this model, and based on many experimental data, the catalytic site is associated with the C-terminal half of the enzyme, although an allosteric site for the binding of glucose 6-phosphate could be present on the N-terminal half of the molecule. We have isolated a cDNA clone of hexokinase from a lambda gt11 human placenta library comprising 2658 bp, containing a single open reading frame of 1893 nucleotides, which encodes a truncate form of hexokinase starting from asparagine-287 to the terminal serine-917. This clone was further digested with restriction enzyme NcoI to obtain almost only the C-terminal half of human hexokinase starting from methionine-455 to the terminal amino acid and was overexpressed in active form in Escherichia coli and purified by ion-exchange h.p.l.c. The overexpressed 'mini'-hexokinase was found not only to catalyse glucose phosphorylation, but also to be inhibited by glucose 6-phosphate and other mono- and bis-phosphate sugars exactly like the complete mammalian enzyme. These results suggest that the C-terminal half of human hexokinase, in addition to the catalytic site, also contains the regulatory site and that the evolutionary relationship between the hexokinases should be reconsidered by including the appearance of a regulatory site before the gene duplication.
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PMID:A recombinant human 'mini'-hexokinase is catalytically active and regulated by hexose 6-phosphates. 163

We show by the use of 32P-labeling in vivo that hexokinase 2 and hexokinase 1 in Saccharomyces cerevisiae are phosphoproteins. The highest labeling was after incubation in medium with a low concentration of glucose, when labeling appears to be predominant even without use of immunoprecipitation. The nature of the modification is not known, but it has properties consistent with a phosphomonoester of serine or threonine. The cAMP-dependent protein kinase plays a negative role in hexokinase phosphorylation, in that there was reduced labeling in strains (bcy1) lacking a regulatory subunit, and increased labeling during growth with high concentrations of glucose in a strain attenuated in the catalytic subunit (tpk1w1). The function of the modification is not known, but there was a correlation between the extent of labeling and the expression of kinase-dependent high-affinity glucose uptake.
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PMID:Phosphorylation of yeast hexokinases. 216 41

The authors described a cooperative effect of glucocorticoids, catecholamines and high density lipoproteins (HDLP), based on increased hexokinase and glucokinase biosynthesis in the liver, resulting in elevated activity of the respective enzymes. This effect was completely blocked by actinomycin-D (a DNA-dependent inhibitor of RNA synthesis), vinblastine (an inhibitor of the microtubular apparatus and intracellular shift) and gordox (an inhibitor of serine proteinases). The modulating effect of HDLP with relation to the adaptive hormones emphasizes the cooperative nature of the new mechanism of enzymatic synthesis induction.
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PMID:[The cooperative effect of hydrocortisone, adrenaline and high-density lipoproteins in regulating liver hexokinase activity]. 236 32

A reactive Glc analog, N-(bromoacetyl)-D-glucosamine (GlcNBrAc), has recently been used (D. M. Schirch and J. E. Wilson (1987) Arch. Biochem. Biophys. 254, 385-396) to label the Glc binding site of rat brain Type I hexokinase. This site has been located in a 40-kDa domain at the C-terminus of the enzyme previously shown to be the location of the substrate ATP binding site (M. Nemat-Gorgani and J. E. Wilson (1986) Arch. Biochem. Biophys. 251, 97-103). In the present study, peptide mapping of hexokinase modified by radiolabeled GlcNBrAc yields three labeled peptides (Peptides I-III). Peptides I and III, as well as catalytic activity, are protected by inclusion of Glc or GlcNAc during reaction with GlcNBrAc. These two peptides show considerable homology to contiguous regions in the sequences of yeast hexokinase isozymes A and B. Peptide III is homologous to a sequence which, based on the X-ray crystallographic work by Steitz and co-workers, is located near the Glc binding site of yeast hexokinase; Peptide I is homologous to an immediately adjacent (toward the C-terminus) region of yeast hexokinase. An essential serine residue implicated in the binding of Glc to the yeast enzyme is also conserved in Peptide III from rat brain hexokinase. These results provide strong support for the view that the "catalytic domain" at the C-terminus of the mammalian Type I hexokinase shares a common ancestry with yeast hexokinase. Peptide II appears to be nonspecifically labeled by GlcNBrAc since labeling is insensitive to the presence of protective ligands such as Glc or GlcNAc; the sequence of Peptide II shows no detectable homology with the yeast isozymes.
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PMID:Rat brain hexokinase: amino acid sequence at the substrate hexose binding site is homologous to that of yeast hexokinase. 363 58

Yeast hexokinase B (ATP:-hexose 6-phosphotransferase, EC 2.7.1.1) was crystallized in the presence of D-xylose and ADP, and its structure was determined at 7 A resolution. The enzyme is in the 'open' conformation which is characteristic of the enzyme crystallized in the absence of glucose, rather than in the 'closed' conformation that is observed with the glucose complex. That is, the binding of xylose into the large cleft that separates the molecule into two lobes does not cause the cleft to close. We conclude, then, that the glucose 6-hydroxymethyl group (which binds to an aspartic acid and a serine) is essential for the hexose-induced conformational change.
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PMID:The 6-hydroxymethyl group of a hexose is essential for the substrate-induced closure of the cleft in hexokinase. 675 1

We recently reported that tyrphostin 23 (3,4-dihydroxybenzylidene malononitrile) is unstable in solution and that some of the degradation products are better inhibitors of the tyrosine kinase activity of Src and the EGF-receptor kinase than the parent compound itself (Ramdas et al., Cancer Res. 54, 867-868, 1994). In this study, the tyrphostin 23-derived compound designated P3, which is a more stable and potent protein tyrosine kinase inhibitor, was isolated. P3 was purified from oxidized tyrphostin 23 by solvent extraction, silica-gel flash chromatography, and reverse-phase high-pressure liquid chromatography. The physical characteristics of the isolated compound were determined and its chemical structure elucidated by 1H and 13C NMR spectroscopy. The proposed structure of this new inhibitor is that of a tyrphostin 23 dimer joined at the benzylidene carbon. P3 was evaluated in vitro as an inhibitor of four different protein tyrosine kinases (Src, Csk, EGF-receptor, and FGF-receptor) and two protein serine kinases (PK-A and PK-C). This compound exhibited the most inhibitory activity against Src with a Ki value of 6 microM and was less inhibitory toward the other protein kinases with Ki values ranging from 35 to 300 microM. P3 did not inhibit other nucleotide-utilizing enzymes such as lactate dehydrogenase and hexokinase. The growth and colony formation of HT-29 colon adenocarcinoma cells that contain activated Src was inhibited by P3 with an IC50 value of approximately 10 microM.
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PMID:A tyrphostin-derived inhibitor of protein tyrosine kinases: isolation and characterization. 748 83

Hexokinase 1 (HK1) purified from rat brain exhibits protein kinase activity, including autophosphorylation and phosphorylation of other protein substrates. The amino acid specificity of rat brain autophosphorylation was analyzed with monoclonal antibodies directed against phosphotyrosine and by acid hydrolysis of the phosphorylated enzyme. The results show that serine, threonine, and tyrosine residues are phosphorylated after incubation with ATP. The stoichiometry of this phosphorylation was 0.2 mole phosphate per mole hexokinase after 30 min of incubation. Evaluation of freshly isolated HK1 with monoclonal anti-phosphotyrosine antibody indicates that the enzyme is phosphorylated at a basal level in its native state. We concluded that rat brain HK1 is a dual specificity protein kinase that is phosphorylated physiologically.
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PMID:Hexokinase autophosphorylation: identification of a new dual specificity protein kinase. 753 90


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