A computer program for fitting and statistically analyzing initial rate data applied to bovine hexokinase type III isozyme.

Abstract Initial rate kinetics were used to study the kinetic mechanism of bovine liver Type III hexokinase (EC 3.7.1.1). The kinetic data were fit to initial rate equations with a computer program written in OMNITAB language. The correct weights to be used in the fitting of the kinetic data were obtained by replicate sample analysis and the effect of various weighting schemes upon the kinetic parameters are shown. The computer program can be changed to evaluate a variety of mathematical models by changing a single command statement in the OMNITAB program. It was concluded from experiments with the dead-end inhibitors for substrates, N -acetylglucosamine and ATP 4− , that the kinetic mechanism is Random Bi Bi. Product inhibition experiments suggest that although the kinetic mechanism is random it may not involve rapid equilibration of enyzme and substrate interaction steps prior to the interconversion of the productive ternary complexes. Bovine liver Type III hexokinase appears to be inhibited by inorganic orthophosphate in the physiological range found in calf liver. This effect appears to be a result of the fact that phosphate is a competitive inhibitor of ATP.

[1]  H. Bachelard,et al.  Cerebral-cortex hexokinase. Elucidation of reaction mechanisms by substrate and dead-end inhibitor kinetic analysis. , 1971, The Biochemical journal.

[2]  T. Ureta,et al.  Multiple molecular forms of ATP:hexose 6-phosphotransferase from rat liver. , 1964, Biochemical and biophysical research communications.

[3]  Robert A. Alberty,et al.  The Relationship between Michaelis Constants, Maximum Velocities and the Equilibrium Constant for an Enzyme-catalyzed Reaction , 1953 .

[4]  D. G. Herries,et al.  The simultaneous estimation of orthophosphate and carbamoyl phosphate and application to the aspartate transcarbamoylase reaction. , 1967, Biochimica et biophysica acta.

[5]  H. Fromm,et al.  Kinetics, mechanism, and regulation of rat skeletal muscle hexokinase. , 1974, Journal of Biological Chemistry.

[6]  F. Ballard.  GLUCOSE UTILIZATION IN MAMMALIAN LIVER. , 1965, Comparative biochemistry and physiology.

[7]  A. Cornish-Bowden,et al.  The direct linear plot. A new graphical procedure for estimating enzyme kinetic parameters. , 1974, The Biochemical journal.

[8]  A. Cornish-Bowden,et al.  Statistical considerations in the estimation of enzyme kinetic parameters by the direct linear plot andother methods. , 1974, The Biochemical journal.

[9]  M. Parry,et al.  Further properties and possibel mechanism of action of adenosine 5'-triphosphate-D-glucose 6-phosphotransferase from rat liver. , 1967, Biochemical Journal.

[10]  H. Fromm,et al.  Kinetic studies of the adenosine 5'-triphosphatase activity of yeast hexokinase and its relationship to the mechanism of action of the enzyme. , 1970, The Journal of biological chemistry.

[11]  M. Parry,et al.  Purification and properties of adenosine 5'-triphospae-D-glucose 6-phosphotransferase from rat liver. , 1966, The Biochemical journal.

[12]  H. Fromm,et al.  Ribitol dehydrogenase. III. Kinetic studies with product inhibition. , 1962, The Journal of biological chemistry.

[13]  Joseph Hilsenrath,et al.  Omnitab, A Computer Program for Statistical and Numerical Analysis , 1970 .

[14]  H. Fromm,et al.  Initial rate studies of adenylosuccinate synthetase with product and competitive inhibitors. , 1969, The Journal of biological chemistry.

[15]  H. Katzen.  The multiple forms of mammalian hexokinase and their significance to the action of insulin. , 1967, Advances in enzyme regulation.

[16]  H. Fromm,et al.  Kinetic studies of the brain hexokinase reaction. , 1962, The Journal of biological chemistry.

[17]  R. Schimke,et al.  Multiple hexokinases of rat tissues. Purification and comparison of soluble forms. , 1966, The Journal of biological chemistry.

[18]  R. Alberty.  On the Determination of Rate Constants for Coenzyme Mechanisms1 , 1958 .

[19]  P. R. Bevington,et al.  Data Reduction and Error Analysis for the Physical Sciences , 1969 .

[20]  H. Fromm,et al.  Kinetic studies of the brain hexokinase reaction. A reinvestigation with the solubilized bovine enzyme. , 1967, Biochemistry.

[21]  U. W. Kenkare,et al.  Bovine brain mitochondrial hexokinase. Solubilization, purification, and role of sulfhydryl residues. , 1972, The Journal of biological chemistry.

[22]  Reinhart Heinrich,et al.  Hexokinase of human erythrocytes. Purification, kinetic model and its application to the conditions in the cell. , 1974, European journal of biochemistry.

[23]  H J Fromm,et al.  Studies on the kinetic mechanism and allosteric nature of bovine brain hexokinase. , 1969, The Journal of biological chemistry.

[24]  H. Fromm,et al.  Kinetic studies of yeast hexokinase. , 1962, The Journal of biological chemistry.

[25]  H. Fromm,et al.  Computer simulation studies with yeast hexokinase and additional evidence for the random Bi Bi mechanism. , 1971, The Journal of biological chemistry.

[26]  R. Basford,et al.  The isolation and purification of solubilized hexokinase from bovine brain. , 1967, Biochemistry.