[Penicillin amidase from E. coli. The determination of the ionization constants of the Michaelis complex ionogenic groups in the enzymatic deacylation reactions of the phenacetyl derivatives of phenylglycine and 7-aminodesacetoxychephalosporanic acid].
pH and temperature relationships of the maximum rate of the reaction of enzymatic hydrolysis of phenacetyl derivatives of phenylglycine (PPG) and 7-aminodesacetoxycephalosporanic acid (7-PADCA) catalysed by immobilized penicillinamidase (IPA) (penicillinamidohydrolase CE 3.5.1.11) were studied. A possibility of applying for the routine methods used in determination of electrochemical properties of ampholites for estimation of the ionization constants of the functional groups of both the enzyme and the enzyme-substrate complex was shown. The applicability of various methods for estimation of the ionization constants is discussed and the ways of determination of the ionization constants and the means of quantitative description of the bell-like pH relationship of the kinetic and equilibrium parameters of the biocatalytic reaction are presented. The equations described were used in analysis of the pH relationship of the immobilized penicillinamidase enzymatic activity in the reactions of 7-PADCA and L-PPG hydrolysis. The estimated ionization constants of the ionogenic groups of the Michaelis complexes were used in quantitative description of the electrochemical state of the complexes at wide pH ranges. The acid properties of the PA and IPA complexes with a number of substrates, such as benzylpenicillin, 7-PADCA, L-PPG and PANAB were compared. The effect of the immobilization procedure, electrochemical properties of the substrates and the reaction products on the electrochemical state of the Michaelis complexes is discussed.
D000215:Acylation; D000581:Amidohydrolases / Q000032:analysis*; D002511:Cephalosporins / Q000494:pharmacology*; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008433:Mathematics; D010399:Penicillin Amidase / Q000032:analysis*; D010648:Phenylacetates / Q000378:metabolism*; D013696:Temperature
D002511:Cephalosporins; D010648:Phenylacetates; C028619:phenylacetyl 7-aminodesacetoxycephalosporanic acid; C016650:phenacetyl-D-alpha aminophenylacetic acid; D000581:Amidohydrolases; D010399:Penicillin Amidase
Penitsillinamidaza iz E. coli. Opredelenie konstant ionizatsii ionogennykh grupp kompleksa Mikhaélisa v reaktsiiakh fermentativnogo deatsilirovaniia fenatsetil'nykh proizvodnykh fenilglitsina i 7-aminodezatsetoksitsefalosporanovoĭ kisloty.