title: Characterization of a novel stable biocatalyst obtained by protein engineering.
abstract: Protein engineering is a powerful tool for the improvement of the properties of biocatalysts. Previously we have applied protein engineering technologies to obtain an extremely stable variant of the thermolysin-like protease from Bacillus stearothermophilus [Van den Burg, Vriend, Veltman, Venema and Eijsink (1998) Proc. Natl. Acad. Sci. U.S.A. 95, 2056-2060]. This variant is much more resistant to denaturing conditions (temperature and denaturing agents) than the wild-type enzyme. An extensive enzymic characterization was undertaken to explore the suitability of the variant as a biocatalyst at high temperatures. By comparing a range of variants with increasing thermal stabilities we show that the additivity of the mutations is accompanied by an increase in activity at elevated temperatures in accordance with the Arrhenius law. The results suggest that the constructed protease variants could be suitable alternatives to proteases that are currently used industrially. Our studies demonstrate how protein engineering can be exploited to obtain high-performance biocatalysts.
authors.affiliation: Department of Genetics, Groningen Biomolecular Sciences and Biotechnology Institute, Kerklaan 30, 9751 NN Haren, The Netherlands.
authors.forename: B
authors.initials: B
authors.lastname: Van den Burg
authors.forename: A
authors.initials: A
authors.lastname: de Kreij
authors.forename: P
authors.initials: P
authors.lastname: Van der Veek
authors.forename: J
authors.initials: J
authors.lastname: Mansfeld
authors.forename: G
authors.initials: G
authors.lastname: Venema
mesh_terms: D002384:Catalysis; D004795:Enzyme Stability; D001411:Geobacillus stearothermophilus / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008666:Metalloendopeptidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D008666:Metalloendopeptidases; C044194:microbial metalloproteinases