title: Substrate as a source of thermodynamic nonideality in enzyme kinetic studies: invertase-catalyzed hydrolysis of sucrose.
abstract: Expressions for the effects of thermodynamic nonideality arising from the use of high concentrations of small substrate in enzyme kinetic studies are derived. Their application to experimental results for the hydrolysis of sucrose by yeast invertase (pH 4.9, 37 degrees C) signifies that the progressive decrease in initial velocity at high sucrose concentration is consistent with the occurrence of isomeric expansion during the transition of an enzyme-substrate complex to its activated state. Ultracentrifuge studies on the yeast enzyme preparation are then used to establish the physical acceptability of the volume change required to account for the kinetic effects in these terms: the postulated expansion of 1.3 liter/mol would represent a mere 0.16% increase in hydrated volume (or a corresponding increase in extent of asymmetry). Finally, although originally interpreted to signify an effect of sucrose on water concentration, published results for the invertase-sucrose system [J. M. Nelson and M. P. Schubert (1928) J. Amer. Chem. Soc. 50, 2188-2193] also find a rational explanation in terms of the present analysis based on effects of thermodynamic nonideality in enzyme kinetic studies.
authors.affiliation: Department of Biochemistry, University of Queensland, St. Lucia, Australia.
authors.forename: K E
authors.initials: KE
authors.lastname: Shearwin
authors.forename: D J
authors.initials: DJ
authors.lastname: Winzor
mesh_terms: D002384:Catalysis; D006026:Glycoside Hydrolases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae; D013395:Sucrose / Q000378:metabolism*; D013816:Thermodynamics; D014461:Ultracentrifugation; D043324:beta-Fructofuranosidase
chemical_list: D046911:Macromolecular Substances; D013395:Sucrose; D006026:Glycoside Hydrolases; D043324:beta-Fructofuranosidase