Abstract
This paper aims to investigate the effects of some salts (NaCl, (NH4)2SO4 and Na2SO4) at pH 5.0, 7.0 and 9.0 on the stability of 13 different immobilized enzymes: five lipases, three proteases, two glycosidases, and one laccase, penicillin G acylase and catalase. The enzymes were immobilized to prevent their aggregation. Lipases were immobilized via interfacial activation on octyl agarose or on glutaraldehyde-amino agarose beads, proteases on glyoxyl agarose or glutaraldehyde-amino agarose beads. The use of high concentrations of salts usually has some effects on enzyme stability, but the intensity and nature of these effects depends on the inactivation pH, nature and concentration of the salt, enzyme and immobilization protocol. The same salt can be a stabilizing or a destabilizing agent for a specific enzyme depending on its concentration, inactivation pH and immobilization protocol. Using lipases, (NH4)2SO4 generally permits the highest stabilities (although this is not a universal rule), but using the other enzymes this salt is in many instances a destabilizing agent. At pH 9.0, it is more likely to find a salt destabilizing effect than at pH 7.0. Results confirm the difficulty of foreseeing the effect of high concentrations of salts in a specific immobilized enzyme.
Highlights
IntroductionThese features make them the best options for the requirements of green chemistry, as they can catalyze a complex process under the mildest experimental conditions [6]
As stated in the introduction section, lipases were immobilized on octyl agarose or glutaraldehyde-amino agarose [111], to have two immobilized preparations via quite different phenomena
Starting with CALA (Figure 1), when the enzyme is immobilized on octyl agarose (Figure 1a–c), and with its inactivation at pH 5.0, the presence of 1 M NaCl presented no effect on enzyme stability, while using 3 M of this salt, the stability was slightly improved
Summary
These features make them the best options for the requirements of green chemistry, as they can catalyze a complex process under the mildest experimental conditions [6] They have evolved to fulfill some physiological requirements (e.g., to give a fast answer under stress situations) and some of their features do not fit those of an industrial biocatalyst: maintain high activity, stability, selectivity and specificity for long periods of time under conditions quite far from the physiological ones and on synthetic substrates. In a very nice example of the use of several techniques, an esterase was supplemented with an additional artificial active center (creating the so-called plurizymes) via enzyme modelling and side-directed mutagenesis [30], its activity was later improved by the same tools [31], and an irreversible inhibitor bearing a catalytic organo-metal complex was designed for one of the active centers and coupled to it, enabling the use of just one enzyme molecule to catalyze a cascade process with an enzyme and a metallic active centers in the same protein molecule [31]
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