Carrier-Free Immobilization of Lipase from <i>Candida rugosa</i> with Polyethyleneimines by Carboxyl-Activated Cross-Linking

Velasco-Lozano, Susana ; López-Gallego, Fernando ; Vázquez-Duhalt, Rafael ; Mateos-Díaz, Juan C. ; Guisán, José M. ; Favela-Torres, Ernesto

Resumen: Carrier-free immobilization of Candida rugosa lipase (CRL) and polymers containing primary amino groups were cross-linked using carbodiimide. To accomplish this, the free carboxyl groups of the enzyme were activated with carbodiimide-succinimide in organic medium, and then the activated proteins were cross-linked with different polyethyleneimines (PEIs). The effect of the cross-linker chain length, the amount of added bovine serum albumin (BSA), and carbodiimide concentration on the catalytic properties of resulting cross-linked enzyme aggregates (CLEAs) was investigated. The CLEAs’ size, shape, specific activity, activity recovery, thermostability and enantioselectivity significantly varied according to the preparation procedure. The highest thermostable CRL-CLEA preparation was obtained with 1.3 kDa polyethyleneimine as cross-linker, 10 mg of BSA and 28 mM of carbodiimide. This preparation is 1.3-fold more active and thermostable than CLEAs prepared by the traditional method of amino cross-linking with glutaraldehyde, and retains 60% of residual activity after 22 h at 50°C. Additionally, the CRL-CLEA preparation showed an enantioselectivity of 91% enantiomeric excess (ee). This immobilization procedure provides an alternative strategy for CLEA production, particularly for enzymes where the traditional method of cross-linking via lysine residues leads to enzyme inactivation.

Idioma: Inglés

DOI: 10.1021/bm500333v

Año: 2014

Publicado en: BIOMACROMOLECULES 15, 5 (2014), 1896-1903
ISSN: 1525-7797

Factor impacto JCR: 5.75 (2014)

Categ. JCR: BIOCHEMISTRY & MOLECULAR BIOLOGY rank: 39 / 290 = 0.134 (2014) - Q1 - T1
Categ. JCR: POLYMER SCIENCE rank: 5 / 82 = 0.061 (2014) - Q1 - T1
Categ. JCR: CHEMISTRY, ORGANIC rank: 5 / 58 = 0.086 (2014) - Q1 - T1

Tipo y forma: Article (PostPrint)