title: [E. coli penicillin amidase. Methods for estimating the close ionization constants of ionogenic groups of the enzyme complex with substrates containing free amino groups].
abstract: The possible use of various procedures for estimation of the ionization constants of the Michaelis complex by the pH dependence of the maximum enzymatic reaction rate is discussed. It is shown that the procedures described in the literature for estimation of the close ionization constants of the enzyme-substrate complexes have limitations and in some cases cannot be used. The paper presents the methods for estimation of the constants and means for quantitative description of the bell-shaped pH dependence of the kinetic and equilibrium parameters of the biocatalytic reaction. The equations recommended in the paper were used in analysis of the pH dependences of the maximum rate of the reactions during the enzymatic synthesis of cefalexin catalysed with immobilized penicillinamidase (IPA) (CE 3.5. 1.11). The ionization constants of the enzyme-substrate complexes of IPA were compared during hydrolysis and synthesis of the compounds acylated with phenylacetic and aminophenylacetic acids. The effect of the nature of the leaving substrate group and added nucleophilic gent on the electrochemical state of the Michaelis complex is discussed.
mesh_terms: D000581:Amidohydrolases / Q000378:metabolism*; D000596:Amino Acids / Q000378:metabolism*; D002506:Cephalexin / Q000138:chemical synthesis; D004926:Escherichia coli / Q000201:enzymology*; D005609:Free Radicals; D005998:Glycine / Q000031:analogs & derivatives / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D066298:In Vitro Techniques; D007700:Kinetics; D008722:Methods; D010399:Penicillin Amidase / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D000596:Amino Acids; D005609:Free Radicals; C033393:methyl phenylglycine; D000581:Amidohydrolases; D010399:Penicillin Amidase; D002506:Cephalexin; D005998:Glycine
vernacular_title: Penitsillinamidaza iz E coli. Metody rascheta blizkikh konstant ionizatsii ionogennykh grupp kompleksa fermenta s substratami, soderzhashchimi svobodnye aminogruppy.