Factors influencing covalent coupling of enzymes on derivatized polyacrylamide gel beads: A survey

Several enzymes (aldolase, aminoacylase, arginase, carboxypeptidase B, chol inesterase, cyclodextrin glycosyltransferase and glucoamylase) were immobil ised by covalent coupling on polyacrylamide bead polymers possessing carbox ylic functional groups activated by water-soluble carbodiimides. The factor s influencing the immobilisation process were studied. The catalytic activi ties of the immobilised enzymes were influenced advantageously by the struc ture of carbodiimide used as coupling agent. Immobilised enzymes of highest catalytic activity could be obtained if the carbodiimide was introduced in to the reaction mixture in a stoichiometric quantity relative to the carbox ylic functional groups located on the support and the support/protein ratio was from 1:0.25 to 1:1. It was found that the hydrogen ion concentration o f the coupling reaction mixture has a profound effect on the immobilisation of enzymes. In the function of the ionic strength of coupling reaction mixture, the cat alytic activity of immobilised enzyme produced showed an apparent optimum. The increasing porosity of support was favourable for the immobilisation of enzymes. The insertion of a spacer in the support appeared to be disadvant ageous for the enzyme immobilisation.