Effect of the Addition of a Nonaqueous Polar Solvent (Glycerol) on Enzymatic Catalysis in Reverse Micelles. Hydrolysis of 2-Naphthyl Acetate by α-Chymotrypsin

Abstract

The kinetics of hydrolysis of 2-naphthyl acetate (2-NA) catalyzed by α-chymotrypsin (α-CT), in reverse micellar solutions formed by glycerol (GY)−water (38% v/v) mixture/sodium bis(2-ethylhexyl)sulfosuccinate (AOT)/n-heptane has been determined by spectroscopic measurements. To compare the efficiency of this reaction with that observed in micelles with water in the core, as well as in the corresponding homogeneous media, the reaction was also studied in water/AOT/n-heptane reverse micellar solutions and in both homogeneous media (water and GY−water, 38% v/v mixture). In every media, α-CT was characterized by the absorption and emission spectra, the fluorescence lifetimes, and the fluorescence anisotropy of its tryptophan residues. The effect of AOT concentration on the kinetic parameters obtained in the micellar systems was determined, at a constant molar ratio of the inner polar solvent and surfactant. Moreover, the data obtained allowed the evaluation of the 2-NA partition constant between the organic and the micellar pseudophase. It is shown that the addition of GY to the micelle interior results in an increase in the catalytic properties of α-CT. The fluorescence anisotropy studies in the different media show that the addition of GY increases the viscosity as compared with the aqueous systems. It seems that the GY addition to the reverse micellar aggregates results in a decrease of the conformational mobility of α-CT, which leads to an increase of the enzyme stability and activity.

On the investigation of the bilayer functionalities of 1,2-di-oleoyl-sn-glycero-3-phosphatidylcholine (DOPC) large unilamellar vesicles using cationic hemicyanines as optical probes: A wavelength-selective fluorescence approach

“…This result in conjunction with the one showed above that the DOPC bilayer is an electron rich microenvironment, seems to us that are quite important specially to take advantage of the DOPC vesicles as "green" nanoreactors systems. This conclusion is based in the well-known capacity that the polar core of the AOT reverse micelles has to create a unique microenvironment for carrying out a variety of chemical and biochemical reactions [54][55][56][57]. Also, they have been used as model systems for studying various reactions in confinement, for example, enzymatic reactions [54,55], or micellar catalysis [56,57].…”

“…This conclusion is based in the well-known capacity that the polar core of the AOT reverse micelles has to create a unique microenvironment for carrying out a variety of chemical and biochemical reactions [54][55][56][57]. Also, they have been used as model systems for studying various reactions in confinement, for example, enzymatic reactions [54,55], or micellar catalysis [56,57]. Indeed, the DOPC bilayer is a "greener" system than the reverse micelles because water is the dispersant medium and DOPC is a nontoxic molecule.…”

The use of acridine orange base (AOB) as molecular probe to characterize nonaqueous AOT reverse micelles

“…The partitioning of AOB between the nonaqueous AOT reverse micelles and the external solvent will be treated within the framework of the pseudophase model [18,38,44,[46][47][48][49][50]. This model considers the microaggregates as a distinct pseudophase whose properties are independent of the AOT concentration and are only determined by the value of the characteristic parameter W s .…”

Distribution of amines in water/AOT/n-hexane reverse micelles: influence of the amine chemical structure

“…According to the pseudophase model, the partition constant K can be defined by [6,[18][19][20]30,31] (2)…”