Immobilization–stabilization of the lipase from Thermomyces lanuginosus: Critical role of chemical amination

• Published in: Process biochemistry (1991) Vol. 44; no. 9; pp. 963 - 968
• Main Authors: Rodrigues, Rafael C., Godoy, Cesar A., Volpato, Giandra, Ayub, Marco A.Z., Fernandez-Lafuente, Roberto, Guisan, Jose M.
• Format: Journal Article
• Language: English
• Published: Elsevier Ltd 01.09.2009
• Subjects: Chemical amination; Enzyme immobilization; Enzyme stabilization; Glyoxyl agarose; Multipoint covalent attachment; Thermomyces lanuginosus; Thermomyces lanuginosus lipase; Glyoxyl agarose; Chemical amination; Enzyme immobilization; Enzyme stabilization; Multipoint covalent attachment; Thermomyces lanuginosus lipase
• ISSN: 1359-5113; 1873-3298
• DOI: 10.1016/j.procbio.2009.04.015

This paper describes the immobilization and stabilization of the lipase from Thermomyces lanuginosus (TLL) on glyoxyl agarose. Enzymes attach to this support only by the reaction between several aldehyde groups of the support and several Lys residues on the external surface of the enzyme molecules at pH 10. However, this standard immobilization procedure is unsuitable for TLL lipase due to the low stability of TLL at pH 10 and its low content on Lys groups that makes that the immobilization process was quite slow. The chemical amination of TLL, after reversible immobilization on hydrophobic supports, has been shown to be a simple and efficient way to improve the multipoint covalent attachment of this enzyme. The modification enriches the enzyme surface in primary amino groups with low p Kb, thus allowing the immobilization of the enzyme at lower pH values. The aminated enzyme was rapidly immobilized at pH 9 and 10, with activities recovery of approximately 70%. The immobilization of the chemically modified enzyme improved its stability by 5-fold when compared to the non-modified enzyme during thermal inactivation and by hundreds of times when the enzyme was inactivated in the presence of organic solvents, being both glyoxyl preparations more stable than the enzyme immobilized on bromocyanogen.