Improvement of chymotrypsin enzyme stability as single enzyme nanoparticles

Abstract

Individual enzyme molecules were coated with a nanometer-scale polymer network in order to increase their stability, providing longer lifetime of enzymes for biochemical processes. This polymer nanolayer is thin and porous enough [Kim, J., Grate, J.W., 2003. Single-enzyme nanoparticles armored by a nanometer-scale organic/inorganic network. Nano Letters 3, 1219–1222] to allow practically unhindered diffusion of the substrate from the solution to the active site of the enzyme, and unhindered transfer of the product from the active site to the solution. During the three-step preparation of the individual enzyme nanoparticles, the chymotrypsin enzyme investigated can lose 30–50% of its original activity. The main cause for a decrease in the enzyme's activity is the UV-light irradiation used during the polymerization step of the pretreatment. The UV-light can destroy the tertiary structure of the enzyme, and this can lead to reduced activity. The enzyme modification and solubilization steps of the treatment methodology do not essentially lower enzyme activity. The morphology and size of enzyme nanoparticles prepared was examined by transmission electron microscopy and demonstrated in this paper. The activity's change of the free and the covered enzyme was investigated at different temperatures, shaking frequencies and pH values. It has been proved that the preparation of enzyme nanoparticles can essentially stabilize the enzyme. Its activity changes much slower than that of native enzyme. The pretreated enzyme can have relatively high residual activity under extreme pH values and temperature, as well. The essence of the results presented is that stability of the enzyme can be increased significantly by covering individual enzymes with a thin, porous polymer layer.