High Stabilization of Enzymes Immobilized on Rigid Hydrophobic Glyoxyl-Supports: Generation of Hydrophilic Environments on Support Surfaces

Catalysts

María Romero-Fernández¹  Alejandro H. Orrego¹  JoséM. Guisán¹  Javier Rocha-Martin¹  Justo Pedroche²  Maríadel Carmen Millán-Linares² 

[1] Department of Biocatalysis, Institute of Catalysis and Petrochemistry (ICP) CSIC, Campus UAM, Cantoblanco, 28049 Madrid, Spain;Group of Plant Proteins, Instituto de la Grasa CSIC, Carretera Utrera Km 1, 41013 Seville, Spain;

关键词: protein stabilization;    protein immobilization;    2-picoline borane;    methacrylic support;    microenvironment;    biocatalysis;   

DOI  :  10.3390/catal10060676

来源: DOAJ

【 摘 要 】

Very rigid supports are useful for enzyme immobilization to design continuous flow reactors and/or to work in non-conventional media. Among them, epoxy-methacrylic supports are easily functionalized with glyoxyl groups, which makes them ideal candidates for enzyme stabilization via multipoint covalent immobilization. However, these supports present highly hydrophobic surfaces, which might promote very undesirable effects on enzyme activity and/or stability. The hydrophilization of the support surface after multipoint enzyme immobilization is proposed here as an alternative to reduce these undesirable effects. The remaining aldehyde groups on the support are modified with aminated hydrophilic small molecules (glycine, lysine or aspartic acid) in the presence of 2-picoline borane. The penicillin G acylase from Escherichia coli (PGA) and alcohol dehydrogenase from Thermus thermophilus HB27 (ADH2) were immobilized on glyoxyl-functionalized agarose, Relizyme and Relisorb. Despite the similar density of aldehyde groups displayed by functionalized supports, their stabilization effects on immobilized enzymes were quite different: up to 300-fold lower by hydrophobic supports than by highly hydrophilic glyoxyl-agarose. A dramatic increase in the protein stabilities was shown when a hydrophilization treatment of the hydrophobic support surface was done. The PGA immobilized on the glyoxyl-Relisorb hydrophilized with aspartic acid becomes 280-fold more stable than without any treatment, and it is even more stable than the PGA immobilized on the glyoxyl agarose.

【 授权许可】

Unknown