Stabilization of penicillin G acylase from Escherichia coli:: Site-directed mutagenesis of the protein surface to increase multipoint covalent attachment

Three mutations on the penicillin acylase surface (increasing the number of Lys in a defined area) were performed. They did not alter the enzyme's stability and kinetic properties; however, after immobilization on glyoxyl-agarose, the mutant enzyme showed improved stability under all tested conditions (e.g., pH 2.5 at 4degreesC, pH 5 at 60degreesC, pH 7 at 55degreesC, or 60% dimethylformamide), with stabilization factors ranging from 4 to 11 compared with the native enzyme immobilized on glyoxyl-agarose.