Abstract
The effect of bio-imprinting and water activity on catalytic activities and the thermostability of lipases was investigated for transesterification using vinyl acetate and benzyl alcohol as substrates in ionic liquid, [Cnmim][PF6] (n=4,6,8), and benzene. The catalytic activities were enhanced by imprinting in benzene and [C4mim][PF6], and the relations between the transesterification activities and the water activity in both solvents were approximately bell shaped. The reactivity of the transesterification in benzene was higher than that in [C4<br /> mim][PF6]. The effects of water activity and imprinting on the kinetic parameters in [C4mim][PF6] were examined. Without controlling the water content, the values of Km,VA and Km,BA (Michaelis constants of vinyl acetate and benzyl alcohol, respectively) decreased, and the values of Vm (maximum rate) increased by imprinting. On the other hand, by controlling the water content in the organic media, the values of Vm, Km,VA, and Km,BA increased by imprinting. The activities of lipase in ionic liquid are more strongly affected by water activity and imprinting than those in benzene. We observed effects of water activity on thermostability but none from imprinting.
Highlights
Enzyme-catalyzed reactions in organic media are widely performed in industrial biotechnology[1]
Prior to the transesterification catalyzed by the imprinted lipase, the effect of the imprinting molecules on the transesterification of vinyl acetate and benzyl alcohol diluted in [C8mim][PF6] was examined
Increase in the alkyl chain length of the carboxylic acid as a substrate analogue reduced the reactivity. This suggests that the preferable conformational change of lipase for binding with vinyl acetate is initially induced by formic acid, and its conformation is retained after removing the formic acid in a process called bio-imprinting[10]
Summary
Enzyme-catalyzed reactions in organic media are widely performed in industrial biotechnology[1]. Many practical applications have been developed in organic media, usage of organic solvents must be reduced in the future[4] because of environmental contamination, physical hazards, toxicity, and volatility circumstances. Alternative media such as supercritical fluids, solvent-free systems, fluorinated solvents and ionic liquids[5] have been proposed. The enzyme does not return to its original conformation due to its rigid structure[10] This method was reported in enzymatic reactions in conventional organic media[11] because the structural memory of bio-imprinted enzyme is lost in aqueous media[12].
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