Hexokinases

A rather curious activating factor which increases the activity of brain, muscle, and yeast hexokinases to different extents has been isolated from erythrocytes of man. The activator is nondialyzable, strongly associated with proteinaceous material, and loses activity on exposure to 80 to 100 C. It does not appear to be a glycolytic enzyme nor does it increase synthesis of ATP, stabilize ATP, or remove inhibitory products. 

It has been postulated that it may participate in the hexokinase reaction in either of two ways (1) by activating the conversion of the hexopy ranose form to hexofuranose, or (2) by mediating a hexokinase-hormone complex. 

Crude muscle hexokinase phosphorylates both glucose and fructose. The two activities may be separated with ammonium sulfate. Crude extracts have a fructose glucose ratio of 0.26 on fractionation with ammonium sulfate between 0.35 and 0.54 saturation, a fructose glucose ratio of 0.148 is observed. Muscle fructokinase is not inhibited by glucose and is saturated by its substrate at unusually high concentrations. The reaction product is not the expected fructose-6-phosphate but rather fructose-1-phosphate. Muscle glucokinase has not as yet been examined to any extent for its specificity. 

Rat liver extracts also contain two highly specific kinases, namely, glucokinase and fructokinase. Glucokinase, in the presence of ATP and Mg++, forms glucose-6-phosphate fructokinase catalyzes the formation of fructose-l-phosphate. Beef Uver fructokinase, like rat fiver fructokinase, is strongly activated by potassium chloride. Sodium and ammonium ions are relatively inert. The affinity of the enzyme for fructose is very high, the Km being lower than 5 X 10 moles per liter. This is in contrast to muscle fructokinase, which has a very weak aflfinity for its subtrate. 

A survey of glucokinase activity in different muscles from different mammals indicates a variation in specific activity. Thus, in the rabbit the sequence of glucokinase activity is heart muscle skeletal muscle stomach muscle diaphragm muscle, the ratio of activities being 1 1.5 0.5 O.2. In the rat the ratio of heart to skeletal muscle activity is 1.4 in the guinea pig, 1.6 and in the chicken, 1.9. Extracts of skeletal muscle of rat and man are significantly more active than those obtained from similar muscles of the guinea pig, rabbit, and chicken. 

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Two or more linked enzyme reactions can lead to a change in the concentration of NADH or NADPH that is equivalent to the concentration of the original analyte. The reference glucose measurement using hexokinase [9001-51-8] and glucose-6-phosphate dehydrogenase [9001-40-5] is an example ... 

Excellent correlation was found when results at 660 nm and 749 nm were compared using a reference hexokinase glucose method (27). The dose response was excellent up to 300 mg/dL glucose. In general, water-borne coatings do not lend themselves to ranging by antioxidants (qv). 

Retention, too, is highly tissue-specific. Sometimes, the extraction mechanism is also the retention mechanism, as for Tc-sestamibi, which is retained in mitochondria as long as transmembrane potentials remain intact. Others are separate. F-2-Fluorodeoxyglucose enters the cell by the same pathway as glucose, but is trapped because it is not a substrate for hexokinase, preventing further intracellular metabohsm. 

Fig. 4. Schematic of a multisequence biosensor in which the target glucose is first converted to glucose-6-phosphate, G6P, in the test solution by hexokinase. G6P then reacts selectively with glucose-6-phosphate dehydrogenase immobilized on the quartz crystal surface. Electrons released in the reaction then chemically reduce the Pmssian blue film (see Fig. 3), forcing an uptake of potassium ions. The resulting mass increase is manifested as a...

ATP 2 ADR The structure was determined to 3.0 A resolution in the laboratory of Georg Schulz in Heidelberg, Germany, (c) The ATP-binding domain of the glycolytic enzyme hexokinase, which catalyzes the phosphorylation of glucose. 

Steitz, T.A., et al. High resolution x-ray structure of yeast hexokinase, an allosteric protein exhibiting a non-symmetric arrangement of subunits. 

Hexokinase catalyzes the phosphorylation of glucose from ATP, yielding glncose-6-P and ADR Using the values of Table 3.3, calculate the standard-state free energy change and equilibrium constant for the hexokinase reaction. 

Mg-" Alcohol dehydrogenase Hexokinase 5 -Deoxyadenosylcobalamin (vitamin Big) H atoms and alkyl groups Me thy Im alony 1-CoA mutase... 

T"he extraordinary ability of an enzyme to catalyze only one particular reaction is a quality known as specificity (Chapter 14). Specificity means an enzyme acts only on a specific substance, its substrate, invariably transforming it into a specific product. That is, an enzyme binds only certain compounds, and then, only a specific reaction ensues. Some enzymes show absolute specificity, catalyzing the transformation of only one specific substrate to yield a unique product. Other enzymes carry out a particular reaction but act on a class of compounds. For example, hexokinase (ATP hexose-6-phosphotransferase) will carry out the ATP-dependent phosphorylation of a number of hexoses at the 6-posi-tion, including glucose. 

Reaction 1 Phosphorylation of Glucose hy Hexokinase or Glucokinase—The First Priming Reaction... 

In most animal, plant, and microbial cells, the enzyme that phosphorylates glucose is hexokinase. Magnesium ion (Mg ) is required for this reaction, as for the other kinase enzymes in the glycolytic pathway. The true substrate for the hexokinase reaction is MgATP. The apparent K , for glucose of the animal... 

In the kidney and in muscle tissues, fructose is readily phosphorylated by hexokinase, which, as pointed out above, can utilize several different hexose substrates. The free energy of hydrolysis of ATP drives the reaction forward ... 

Another simple sugar that enters glycolysis at the same point as fructose is mannose, which occurs in many glycoproteins, glycolipids, and polysaccharides (Chapter 7). Mannose is also phosphorylated from ATP by hexokinase, and the mannose-6-phosphate thus produced is converted to fructose-6-phosphate by phosphomannoisomerase. 

Yet another difference is that laboratory reactions are often done using relatively small, simple reagents such as Br2, HC1, NaBH4, OO3, and so forth, while biological reactions usually involve relatively complex "reagents" called coenzymes. In the hexokinase-catalyzed phosphorylation of glucose just shown,... 

Acifluorfen, synthesis of, 683 Acrolein, structure of, 697 Acrylic acid, pKa of, 756 structure of. 753 Activating group (aromatic substitution), 561 acidity and, 760 explanation of, 564-565 Activation energy, 158 magnitude of, 159 reaction rate and, 158-159 Active site (enzyme), 162-163 citrate synthase and, 1046 hexokinase and, 163... 

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