Probing Immobilization Mechanism of alpha-chymotrypsin onto Carbon Nanotube in Organic Media by Molecular Dynamics Simulation

Abstract

The enzyme immobilization has been adopted to enhance the activity and stability of enzymes in non-aqueous enzymatic catalysis. However, the activation and stabilization mechanism has been poorly understood on experiments. Thus, we used molecular dynamics simulation to study the adsorption of a-chymotrypsin (a-ChT) on carbon nanotube (CNT) in aqueous solution and heptane media. The results indicate that a-ChT has stronger affinity with CNT in aqueous solution than in heptane media, as confirmed by more adsorption atoms, larger contact area and higher binding free energies. Although the immobilization causes significant structure deviations from the crystal one, no significant changes in secondary structure of the enzyme upon adsorption are observed in the two media. Different from aqueous solution, the stabilization effects on some local regions far from the surface of CNT were observed in heptane media, in particular for S1 pocket, which should contribute to the preservation of specificity reported by experiments. Also, CNT displays to some extent stabilization role in retaining the catalytic H-bond network of the active site in heptane media, which should be associated with the enhanced activity of enzymes. The observations from the work can provide valuable information for improving the catalytic properties of enzymes in non-aqueous media.O ver the past two decades, non-aqueous enzymatic catalysis has attracted considerable interests from experiments and theories since it can provide numerous synthetic and processing advantages compared to aqueous environment, for example, higher selectivity, thermo-stability, lower side reactions 1-3 . Thus, it could allow the syntheses of some compounds that are difficult to obtain in aqueous solution. Nevertheless, the enzyme in non-aqueous solution exhibits very low activity and lacks of long-term operational stability, which significantly limited its applications in industry. To tackle these problems, considerable experimental efforts have been devoted to activate enzymes in non-aqueous media, for example, salt activation, chemical modification and enzyme immobilization 4 . As known, the immobilization has been prevalently used in the classical enzymatic catalysis in aqueous media to improve the catalytic properties of enzymes [5][6][7][8] . Also, the method was adopted in the enzymatic catalysis in non-aqueous media, and the enhanced enzyme activity, stability and recyclability 4,9-12 were reported. For example, glass-adsorbed chymotrypsin exhibited much higher activity than the suspended enzyme (by 1-2 orders of magnitude) in some polar and nonpolar organic media 10 . Subtilisin Carlsberg and a-chymotrypsin adsorbed onto silica support led to a significant increase in their activity in acetonitrile media with respect to the native enzyme preparation 11 . Lipases immobilized on poly(acrylonitrile-co-maleic acid) ultrafiltration hollow fiber membrane were observed to have higher hydrolytic activity and the operational stability than its free ones in heptane media 12 . Fo...

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