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 4 degrees C, pH 5 at 60 degrees C, pH 7 at 55 degrees C, or 60% dimethylformamide), with stabilization factors ranging from 4 to 11 compared with the native enzyme immobilized on glyoxyl-agarose.
Abian
Olga
O
Departamento de Biocatalisis Instituto de Catálisis y Petroleoquímica, CSIC, Universidad Autónoma de Madrid, 28049 Madrid, Spain.
Grazú
Valeria
V
Hermoso
Juan
J
González
Ramón
R
García
José Luis
JL
Fernández-Lafuente
Roberto
R
Guisán
José Manuel
JM
D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010399:Penicillin Amidase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D010399:Penicillin Amidase