title: Kinetic studies with myo-inositol monophosphatase from bovine brain.
abstract: The kinetic properties of myo-inositol monophosphatase with different substrates were examined with respect to inhibition by fluoride, activation or inhibition by metal ions, pH profiles, and solvent isotope effects. F- is a competitive inhibitor versus 2'-AMP and glycerol 2-phosphate, but noncompetitive (Kis = Kii) versus DL-inositol 1-phosphate, all with Ki values of approximately 45 microM. Activation by Mg2+ follows sigmoid kinetics with Hill constants around 1.9, and random binding of substrate and metal ion. At high concentrations, Mg2+ acts as an uncompetitive inhibitor (Ki = 4.0 mM with DL-inositol 1-phosphate at pH 8.0 and 37 degrees C). Activation and inhibition constants, and consequently the optimal concentration of Mg2+, vary considerably with substrate structure and pH. Uncompetitive inhibition by Li+ and Mg2+ is mutually exclusive, suggesting a common binding site. Lithium binding decreases at low pH with a pK value of 6.4, and at high pH with a pK of 8.9, whereas magnesium inhibition depends on deprotonation with a pK of 8.3. The pH dependence of V suggests that two groups with pK values around 6.5 have to be deprotonated for catalysis. Solvent isotope effects on V and V/Km are greater than 2 and 1, respectively, regardless of the substrate, and proton inventories are linear. These results are consistent with a model where low concentrations of Mg2+ activate the enzyme by stabilizing the pentacoordinate phosphate intermediate. Li+ as well as Mg2+ at inhibiting concentrations bind to an additional site in the enzyme-substrate complex. Hydrolysis of the phosphate ester is rate limiting and facilitated by acid-base catalysis.
authors.affiliation: Merrell Dow Research Institute, Strasbourg, France.
authors.forename: A J
authors.initials: AJ
authors.lastname: Ganzhorn
authors.forename: M C
authors.initials: MC
authors.lastname: Chanal
mesh_terms: D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000201:enzymology*; D002417:Cattle; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D012969:Sodium Fluoride / Q000494:pharmacology; D012997:Solvents
chemical_list: D012997:Solvents; D012969:Sodium Fluoride; D010744:Phosphoric Monoester Hydrolases; C021822:myo-inositol-1 (or 4)-monophosphatase; D008274:Magnesium