Radiation-induced polymerization for the immobilization of penicillin acylase.
The immobilization of Escherichia coli penicillin acylase (EC 3.5.1.11) was investigated by radiation-induced polymerization of 2-hydroxyethyl methacrylate at low temperature. A leak-proof composite that does not swell in water was obtained by adding the cross-linking agent trimethylolpropane trimethacrylate to the monomer-aqueous enzyme mixture. Penicillin acylase, which was immobilized with greater than 70% yield, possessed a higher Km value toward the substrate 6-nitro-3-phenylacetamidobenzoic acid than the free enzyme form (Km = 1.7 X 10(-5) and 1 X 10(-5) M, respectively). The structural stability of immobilized penicillin acylase, as assessed by heat, guanidinium chloride, and pH denaturation profiles, was very similar to that of the free-enzyme form, thus suggesting that penicillin acylase was entrapped in its native state into aqueous free spaces of the polymer matrix.
Boccù
E
E
Dipartimento di Scienze Farmaceutiche, Università di Padova, Italy.
Carenza
M
M
Lora
S
S
Palma
G
G
Veronese
F M
FM
D000581:Amidohydrolases / Q000528:radiation effects*; D004800:Enzymes, Immobilized / Q000378:metabolism / Q000528:radiation effects*; D004926:Escherichia coli / Q000201:enzymology; D005720:Gamma Rays; D007700:Kinetics; D046911:Macromolecular Substances; D008689:Methacrylates / Q000528:radiation effects; D010399:Penicillin Amidase / Q000378:metabolism / Q000528:radiation effects*
D004800:Enzymes, Immobilized; D046911:Macromolecular Substances; D008689:Methacrylates; C005044:hydroxyethyl methacrylate; D000581:Amidohydrolases; D010399:Penicillin Amidase; C028643:trimethylolpropane trimethacrylate