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 Previous Article

Applied and Environmental Microbiology, February 2004, p. 1249-1251, Vol. 70, No. 2
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.2.1249-1251.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Stabilization of Penicillin G Acylase from Escherichia coli: Site-Directed Mutagenesis of the Protein Surface To Increase Multipoint Covalent Attachment

Olga Abian,1 Valeria Grazú,1 Juan Hermoso,2 Ramón González,3 José Luis García,4 Roberto Fernández-Lafuente,1* and José Manuel Guisán1*

Departamento de Biocatalisis Instituto de Catálisis y Petroleoquímica, CSIC, Universidad Autónoma de Madrid, 28049 Madrid,1 Instituto de Química-Física Rocasolano,2 Instituto de Fermentaciones Industriales, CSIC, 28006 Madrid,3 Centro de Investigaciones Biológicas, CSIC, 28008 Madrid, Spain4

Received 6 October 2003/ Accepted 13 November 2003

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°C, pH 5 at 60°C, pH 7 at 55°C, or 60% dimethylformamide), with stabilization factors ranging from 4 to 11 compared with the native enzyme immobilized on glyoxyl-agarose.

* Corresponding author. Mailing address: Departamento de Biocatalisis, Instituto do Catálisis y Petroleoquímica, CSIC, Universidad Autónoma de Madrid, Cantoblanco, 28049 Madrid, Spain. Phone: 34 91 585 48 09. Fax: 34 91 585 47 60. E-mail for Roberto Fernández-Lafuente: rfl{at}icp.csic.es. E-mail for José Manuel Guisán: jmguisan{at}icp.csic.es.

Applied and Environmental Microbiology, February 2004, p. 1249-1251, Vol. 70, No. 2
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.2.1249-1251.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Kim, C.-S., Pierre, B., Ostermeier, M., Looger, L. L., Kim, J. R. (2009). Enzyme stabilization by domain insertion into a thermophilic protein. Protein Eng Des Sel 22: 615-623 [Abstract] [Full Text]  
  • Montes, T., Grazu, V., Lopez-Gallego, F., Hermoso, J. A., Garcia, J. L., Manso, I., Galan, B., Gonzalez, R., Fernandez-Lafuente, R., Guisan, J. M. (2007). Genetic Modification of the Penicillin G Acylase Surface To Improve Its Reversible Immobilization on Ionic Exchangers. Appl. Environ. Microbiol. 73: 312-319 [Abstract] [Full Text]  
  • Scaramozzino, F., Estruch, I., Rossolillo, P., Terreni, M., Albertini, A. M. (2005). Improvement of Catalytic Properties of Escherichia coli Penicillin G Acylase Immobilized on Glyoxyl Agarose by Addition of a Six-Amino-Acid Tag. Appl. Environ. Microbiol. 71: 8937-8940 [Abstract] [Full Text]  


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