Conjugation with inulin improves the environmental stability of haloalkane dehalogenase DhaA

Haloalkane dehalogenase DhaA catalyzes the hydrolytic cleavage of carbon-halogen bonds and produces alcohol, a proton and a halide. However, DhaA suffers from the poor environmental stability, such as sensitivity to high temperature, low pH, hypersaline and organic solvent. In order to improve the environmental stability of DhaA, DhaA was covalently conjugated with inulin, a hydrophilic polysaccharide in the present study. Each DhaA was averagely conjugated with 7-8 inulin molecules. The conjugated inulin could form a hydration layer around DhaA, which increased the conformational rigidity and decreased the entropy of the enzyme. Conjugation of inulin maintained 75.5 % of the enzymatic activity of DhaA and slightly altered the structure of DhaA. As compared with DhaA, the conjugate (inu-DhaA) showed slightly different kinetic parameters (Km of 2.9 mu mol/L and Kcat of 1.0 s-1). Inulin conjugation could delay the structural unfolding and/or slow the protonation process of DhaA under undesirable environment, including the long-term storage, low pH, hypersaline and organic solvent stability. As a result, the environmental stability of DhaA was markedly increased upon conjugation with inulin. Thus, inulin conjugation was an effective approach to enhance the environmental stability of DhaA.

Automated summary

In this study, haloalkane dehalogenase DhaA was covalently conjugated with inulin, a hydrophilic polysaccharide, to enhance the environmental stability of the enzyme. The conjugated inulin formed a hydration layer around DhaA, increasing conformational rigidity and decreasing enzyme entropy. Despite slight changes in structure and enzymatic activity, inulin conjugation delayed structural unfolding and protonation of DhaA in unfavorable conditions. As a result, the environmental stability of DhaA was significantly improved.