Enzymatic Attack on Immobilized Substrates

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Abstract
The chymotrypsin(EC 3.4.21.1)-catalyzed hydrolysis of polyacrylamide-bound L-phenylalanine 4-nitroanilide was studied. As a spacer, 1 or 2 6-aminohexanoyl residues were inserted between the matrix and ligand. In the course of the enzymatic hydrolysis of polyacrylamide-bound substrates, enzyme adsorption by the gel substrates was observed. A quasi equilibrium of enzyme partitioning was reached after approximately 20 min incubation time. The enzyme adsorption could be described by the Langmuir adsorption isotherm. The substrates attached via spacers to the matrix were completely hydrolyzed. The initial course of the product vs. time curves, as well as the dependence of the initial hydrolysis rates on enzyme concentration or substrate concentration, were interpreted by the Nernst reaction theory. The initial rate of the hydrolysis of polyacrylamide-bound L-phenylalanine 4-nitroanilide probably depends on the velocity of enzyme diffusion into the matrix.