Cationic Reverse Micelles Create Water with Super Hydrogen‐Bond‐Donor Capacity for Enzymatic Catalysis: Hydrolysis of 2‐Naphthyl Acetate by α‐Chymotrypsin

Abstract

The reverse micelles (RMs) are very good as nanoreactors because they can create a unique microenvironment for carrying out a variety of chemical and biochemical reactions. The aim of the present work is to determine the influence of different RMs interfaces on the hydrolysis of 2-naphthyl acetate (2-NA) by a-Chymotrypsin (a-CT), The reaction was studied in water/benzyl-n-hexadecyldimethylammonium chloride (BHDC)/benzene RMs and, compare its efficiency with that observed in pure water and in sodium 1,4-bis-2-ethylhexylsulfosuccinate (AOT) RMs. Hence, the hydrolysis rates of 2-NA catalyzed by a-CT have been determined by spectroscopic measurements. In addition, the method used allows the joint evaluation of the substrate partition constant, Kp, between the organic and the micellar pseudophase and the kinetic parameters: catalytic rate constant, kcat, and the Michaelis constant, KM, of the enzymatic reaction. The effect of the surfactant concentration on the kinetics parameters was determined at constant W0 = [H2O]/[Surfactant] and, also the variation of W0 at surfactant concentration constant was investigated. The results show that the classical Michaelis ¡V Menten mechanism is valid for a-CT in all the RMs systems studied and, that the reaction takes place at both RM interfaces. Moreover, the catalytic efficiency kcat/KM values obtained in the RMs systems are higher than the value reported in water. Furthermore, there is a remarkable increase in the a-CT efficiency in the cationic RMs in comparison with the anionic system believably due to the unique water properties found in these confined media. The results show that in cationic RMs the hydrogen bond donor capacity of the water is enhanced due to its interaction with the cationic interface. Hence, water entrapped can be converted into ¡§super-water¡¨ for the enzymatic reaction studied in this work.