engleski

Lyophilization conditions for the stabilization of halohydrin dehalogenases

Halohydrin dehalogenases (HHDHs) catalyse enantioselective formation and conversion of epoxides. Their exceptional selectivity, regioselectivity, and ability to employ different nucleophiles make the enzymes attractive for biocatalysis.[1] The enzyme from Agrobacterium radiobacter AD1 (HheC) is usually expressed in E.coli, isolated in TEMG buffer (50 mM Tris-SO4, 10 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.[2] Both storage (before use) and operational stability (during use) are highly relevant for biotechnological applications.[3] Lyophilization is an attractive approach for the longterm storage of enzymes at ambient temperature. It also facilitates the use of enzymes in non-aqueous media.[4] In this work, the storage stability of HheC isolated in TEMG and TEM buffer is evaluated at room temperature. With a focus on establishing the optimal lyophilized formulation, different additives were tested for enzyme stabilization during lyophilization. Optimal conditions for the lyophilization of several HHDHs have been investigated. 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 (Figure 1).

halohydrin dehalogenase ; enzyme stability ; lyophilization

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