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Activity and stability of lyophilized halohydrin dehalogenase

Stability of enzymes has a significant impact on their use in an alternative reaction media. Presence of organic solvents or exposure to high temperatures leads to denaturation of enzymes since they have evolved to work in the cellular environment. Lyophilization is as an attractive solution for long-term storage of enzymes at ambient temperature. Also, it can be used as a method of enzyme stabilization for their application in non-aqueous media. Halohydrin dehalogeneses are enzymes that catalyse the formation and conversion of epoxides. Reactions are catalyzed in regioselective and enantioselective manner, making the enzymes attractive for biocatalysis. Enzyme from Agrobacterium radiobacter AD1 (HheC) is usually expressed in E.coli, isolated in TEMG buffer (50 mM Tris-SO4, 2 mM EDTA, 10 % glycerol and 1 mM mercaptoethanol), and usually is stored as a cell- free extract at -70 °C for at least three months without significant loss of activity. In this work, the stability of HheC is first evaluated in different buffers (TEMG, TEM and Tris-SO4) at different temperatures. With the aim of obtaining the optimal lyophilized formulation, different additives were tested for HheC stabilization. Lyophilized HheC was incubated at 50 °C to intensify the differences induced by additives. Enzyme activity is determined by following the absorbance at 310 nm.

halohydrin dehalogenase ; protein stability ; lyophilization

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