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)

We describe a new, nonenzymatic method for determining serum glucose. This method is based upon the direct electrochemical oxidation of glucose by means of stable nickel-catalyst that is electrodeposited onto a lead dioxide electrode surface. Linear calibration plots are obtained for glucose in the 0.1 to 4500 mg/liter range. Values for 23 sera with glucose concentrations ranging from 300 to 4300 mg/liter showed a linear correlation coefficient of 0.996 when compared with values reported by a hospital laboratory using the hexokinase method. Typically a 25-microliter sample is used, but samples as small as 5 microliter have been successfully analyzed. The sample is added to 25.00 ml of pH 13 electrolyte containing 1 mmol of NiSO4 per liter; the amount of increase in anodic current comproses the analytical signal. For 20 repetitive analyses of a serum specimen, the CV was less than 4%. Ascorbic acid is rapidly decomposed in the electrolyte used and thus does not interfere. Uric acid is inactive at the concentrations present in serum samples. The sensitivity and simplicity of the new method suggest it as an alternative to currently available procedures.
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PMID:Amperometric nonenzymatic determination of serum glucose by means of a nickel-catalyst electrode. 63 Jul 6

The effects of monovalent (Li+, Cs+) divalent (Cu2+, Ca2+, Sr2+, Ba2+, Zn2+, Cd2+, Hg2+, Pb2+, Mn2+, Fe2+, Co2+, Ni2+) and trivalent (Cr3+, Fe3+, Al3+) metals ions on hexokinase activity in rat brain cytosol were compared at 500 microM. The rank order of their potency as inhibitors of brain hexokinase was: Cr3+ (IC50 = 1.3 microM) greater than Hg2+ = Al3+ greater than Cu2+ greater than Pb2+ (IC50 = 80 microM) greater than Fe3+ (IC50 = 250 microM) greater than Cd2+ (IC50 = 540 microM) greater than Zn2+ (IC50 = 560 microM). However, at 500 microM Co2+ slightly stimulated brain hexokinase whereas the other metal ions were without effect. That inhibition of brain glucose metabolism may be an important mechanism in the neurotoxicity of metals is suggested.
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PMID:Differential effects of monovalent, divalent and trivalent metal ions on rat brain hexokinase. 286 Oct 11

Bivalent metal ions, particularly Zn2+ and other members of the first-row transition series, promote irreversible inactivation of yeast hexokinase by Cibacron Blue F3G-A at a site competitive with both ATP and D-glucose. Difference spectroscopy indicates that the protein-dye dissociation constant is decreased from 250 micrometers in the absence of metal ions to less than 100 micrometers in the presence of appropriate concentrations of metal ions, with specificity displayed in the sequence of Zn2+ greater than Cu2+ greater than Ni2+ greater than Mn2+. Quantitative inactivation of yeast hexokinase leads to the incorporation of approx. 1 mol of Cibacron Blue F3G-A/mol of subunit of mol. wt. 51 000 in both the presence and the absence of metal ion. These results suggest the formation of a highly specific ternary complex involving enzyme, dye and metal ion at the active-site region of the enzyme, and correlate well with the known effects of metal ions in promoting the binding of hexokinase to immobilized Cibacron Blue F3G-A.
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PMID:Metal-ion-promoted binding of triazine dyes to proteins. The interaction of Cibacron Blue F3G-A with yeast hexokinase. 675 35

Low concentrations of metal ions, particularly those of the first row transition series such as Zn2+, Co2+, Mn2+, Ni2+, Cu2+, and, to a lesser extent, the group IIA ions, Ca2+ and Mg2+, promotes binding of carboxypeptidase G2, alkaline phosphatase and yeast hexokinase to immobilized Procion Red H-8BN, Procion Yellow H-A and Cibacron Blue F3G-A respectively. The binding of ovalbumin to immobilized Cibacron Blue F3G-A and Procion Orange MX-G is selectively enhanced in the presence of AI3+. With ovalbumin and alkaline phosphatase, the effect is almost totally specific for both the metal ion and dye, whereas with carboxypeptidase G2 and hexokinase, metal ions such as Co2+, Ni2+, Mn2+, Cu2+, Ca2+ and Mg2+ also promote binding to varying degrees. Almost all other monovalent and trivalent metal ions appear to be ineffective. Metal ion-bound enzymes can subsequently be eluted with appropriate chelating agents of the amine, aminocarboxylate or substituted pyridine classes.
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PMID:Metal ion-promoted binding of proteins to immobilized triazine dye affinity adsorbents. 689 1

Inhibition by its product, glucose, is a kinetic property of hexokinase type III. In this paper, we report the overexpression in Escherichia coli of human hexokinase type III. The recombinant enzyme was genetically fused with a hexahistidine peptide at the C-terminal end. This modification confers to the product the ability to bind the Ni2+ ion immobilised into agarose by nitrilotriacetic acid (NTA) groups. The purification was performed by one-step column chromatography using ammonium sulphate as stabilising agent. Recombinant hexokinase type III appears as a single band of approximately 100 kDa on a SDS-PAGE gel and shows specific activity of 16 U/mg. Its kinetic parameters are comparable to those of the native enzyme, including the fact that it can be inhibited by glucose. The comparison of these results with the properties of the overexpressed carboxyl-domain led us to suppose that the inhibition site for glucose required the presence of the N-terminal domain.
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PMID:The overexpressed hexahistidine-tagged human hexokinase type III is inhibited by D-glucose. 1245 31

The interaction between nickel and yeast hexokinase was studied. The binding of nickel showed a positive cooperativity, and saturation was not reached. The nickel binding induced modifications in the secondary structure of the protein; thus, a lost of alpha helix and beta turns, as well as an increase of the random structure and beta sheet was observed. The monomer/dimmer equilibrium of the protein was modified in the presence of nickel, and the monomer state was mainly obtained at the highest nickel concentrations studied. These changes on the protein structure caused a decrease in the enzyme activity. According to kinetic studies, nickel caused a non-competitive inhibition when glucose was the variable substrate and a linear competitive inhibition when ATP was the variable substrate.
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PMID:Structural and functional implications of the hexokinase-nickel interaction. 1625 2

The full encoding sequence for human type II hexokinase (HXK II) was cloned into the E. coli expression vector pET 21b and expressed as a C-terminally hexahistidine-tagged protein in the BL21 (DE3) strain. The IPTG-induced HXK II approximately accounted for 17% of the total E. coli proteins, and 81% of HXK II(6xHis) existed in inclusion bodies. To improve the production of soluble recombinant HXK II protein, in the functionally active form, we used low temperature, and the osmotic stress expression method. When expressed at 18 degrees C, about 83% of HXK II(6xHis) existed in the soluble fraction, which amounted to a 4.1-fold yield over that expressed at 37 degrees C. The soluble form of HXK II(6xHis) was also highly produced in the presence of 1 M sorbitol under the standard condition (37 degrees C), which indicated that temperature downshift and low water potentials were required to improve the yield of active recombinant HXK II protein. The expressed protein was purified by metal chelate affinity chromatography performed in an IDA Excellose column charged with Ni2+ ions, resulting in about 40 mg recombinant HXK II protein obtained with purity over 89% from 5 l of E. coli culture. The identity of HXK II(6xHis) was confirmed by Western blotting analysis. Taken together, using the stress-governed expression described in this study, human active HXK II can be purified in sufficient amounts for biochemical and biomedical studies.
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PMID:Stress-governed expression and purification of human type II hexokinase in Escherichia coli. 1805 Dec 76

This paper reports a flow-injection analysis (FIA) of glucose not using enzyme based on the electrocatalytic oxidation of glucose at a nickel electrode. The electrocatalytic mechanism and quantificational method of glucose have been investigated. The current intensity of the electrocatalytic oxidation to glucose at the potential of 550mV is proportional to the concentration of glucose over the range of 0.10-2.50mmoll(-1), with a 0.04mmoll(-1) detection limit (S/N=3) and a correlation coefficient of 0.9991. The relative standard deviation (R.S.D.) is less than 4.3% (n=5) for the determination of practical serum samples. The biologic compounds probably existed in the sample, such as ascorbic acid, uric acid, dopamine and epinephrine, do not disturb the determination of glucose. The result is satisfactory for the determination of glucose in human serum sample as comparison to that from the routine hexokinase method.
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PMID:Flow-injection analysis of glucose without enzyme based on electrocatalytic oxidation of glucose at a nickel electrode. 1907 21