Aldehyde-agarose gels as activated supports for immobilization-stabilization of enzymes

Abstract

Aldehyde groups, moderately separated from support surfaces, are proposed as suitable active groups for developing strategies to insolubilize-stabilize enzymes by multipoint covalent attachment to activated preexistent supports. A method for preparation of glyoxyl-Sepharose CL gels, Ag-O-CH 2-CHO, with very different surface densities of active groups, up to 17 aldehyde residues per 1000 Å 2 of gel surface, is presented. These activated gels are very stable, even in moderately alkaline media, e.g., half-life time of these aldehyde groups at pH 10, 25°C, is 12 days. Experiments of insolubilization of the enzyme Penicillin G acylase on these supports indicate that one point amine-aldehyde attachments are fast but quite reversible. However, the binding of the enzyme on the support can be dramatically stabilized by two-point enzyme-support attachment. These qualities, as well as the absence of steric hindrance for the amine-aldehyde chemical reaction, make this activation method very suitable for designing intense, but not distorting, enzyme-support multiinteraction processes.