Thermostability of Cromobacterium viscosum lipase in AOT/isooctane reverse micelle

Abstract

The thermostability of Cromobacterium viscosum lipase (EC 3.1.1.3) entrapped in AOT (sodium bis-[2-ethylhexyl] sulfosuccinate) reverse micelles was increased by the addition of short-chain polyethylene glycol (PEG 400). Two different approaches were considered: (1) the determination of half-life time and (2) the mechanistic analysis of deactivation kinetics. The half-life of lipase entrapped in AOT/isooctane reverse micelles with PEG 400 at 60 degrees C was 28 h, ninefold higher than that in reverse micelles without PEG 400. The lipase entrapped in both reverse micellar systems followed a series-type deactivation mechanism involving two first-order steps. The deactivation constant for the first step at 60 degrees C in PEG containing reverse micelles was 0.055 h!1, 11-fold lower than that in reverse micelles without PEG, whereas it remained almost constant for the second step. The inactivation energy of the lipase entrapped in reverse micelles with and without PEG 400 was 88.12 and 21.97 kJ/mol, respectively.

Thermal Deactivation Kinetics of Pseudomonas fluorescens Lipase Entrapped in AOT/Isooctane Reverse Micelles

“…The data for deactivation in the 10 mM phosphate buffer (pH 8.0), glycerol, or AOT/isooctane reverse micelles were fitted to a two-step series-type deactivation kinetic model, and the kinetic parameters were determined using a nonlinear regression procedure based on the Marquardt–Levenberg method of iterative convergence included in a solver tool of Microsoft Office Excel 2007 software.…”

“…After incubation of the enzyme dissolved in 10 mM phosphate buffer (pH 8.0), glycerol, or reverse micelles consisting of 50 mM AOT-isooctaneglycerol pool at 40, 50, 60, and 70 °C, aliquots were taken at the indicated times, and the residual glycerolysis activity was assayed according to the procedure described in the previous section Determination of the Lipase Deactivation Kinetic Parameters. The data for deactivation in the 10 mM phosphate buffer (pH 8.0), glycerol, or AOT/isooctane reverse micelles were fitted to a twostep series-type deactivation kinetic model, 18 and the kinetic parameters were determined using a nonlinear regression procedure based on the Marquardt−Levenberg method of iterative convergence included in a solver tool of Microsoft Office Excel 2007 software.…”

AOT/isooctane reverse micelles with a microaqueous core act as protective shells for enhancing the thermal stability of Chromobacterium viscosum lipase

“…The data for the deactivation in the 50 mM Tris-HCl buffer (pH 8.0), glycerol (used for glycerolysis), and AOT/isooctane reverse micelles were fitted to a two-step series-type deactivation kinetic model (Knezevic, Siler-Marinkovic, & Mojovic, 1998;Moquin & Temelli, 2008;Talukder, Zaman, Hayashi, Wu, & Kawanishi, 2007). The kinetic parameters were determined using a non-linear regression procedure based on the Marquardt-Levenberg method of iterative convergence included in a solver tool in Microsoft Office Excel 2007 (Microsoft, Redmond, WA, USA) (Valério et al, 2009).…”