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RevistaInternational Journal of Biological Macromolecules
Año2020
Volumen145
Páginas856-864
Internacional

COIMMOBILIZATION OF DIFFERENT LIPASES: SIMPLE LAYER BY LAYER ENZYME SPATIAL ORDERING

Autores:Roberto Fernandez-Lafuente
Grupos de investigación:Optimización de biocatalizadores y bioprocesos enzimáticos
Sara Arana-Peña1,+, Nathalia S. Rios1,2,+, Carmen Mendez-Sanchez1, Yuliya Lokha1,  Diego  Carballares1, Luciana R. B. Gonçalves2 and Roberto Fernandez-Lafuente1,*
1Departamento de Biocatálisis. ICP-CSIC, Campus UAM-CSIC, Madrid, Spain.
2Departamento de Engenharia Química, Universidade Federal do Ceará, Campus do Pici, Bloco 709, CEP 60455-760, Fortaleza, CE, Brazil





This paper shows the step by step coimmobilization of up to five different enzymes following two different orders in the coimmobilization to alter the effect of substrate diffusion limitations. The enzymes were the lipases A and B from Candida antarctica, the lipases from Rhizomocur miehei and, Themomyces lanuginosus and the phospholipase Lecitase Ultra. The utilized strategy was a layer by layer immobilization, coating the immobilized enzymes with polyethylenimine followed by the crosslinking of the enzyme and PEI with glutaraldehyde to prevent enzyme release, and them adding a new lipase layer. The use of previously inactivated biocatalysts (using diethyl p-nitrophenylphosphate) permitted to visualize the immobilization of each enzyme layer, which was later confirmed by SDS-PAGE. This also confirmed the successful and complete covalent crosslinking of the glutaraldehyde treated enzyme layers. Activity of the combibiocatalysts was followed using diverse substrates. The protocol was successful and permitted to immobilize in an ordered way the 5 different enzymes in a down-up distribution.

Palabras clave:enzyme coimmobilization; combilipases; enzyme ordering
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