Crystallization and Crystallographic Analysis of a <i>Bradyrhizobium Elkanii</i> USDA94 Haloalkane Dehalogenase Variant with an Eliminated Halide-Binding Site
oleh: Tatyana Prudnikova, Barbora Kascakova, Jeroen R. Mesters, Pavel Grinkevich, Petra Havlickova, Andrii Mazur, Anastasiia Shaposhnikova, Radka Chaloupkova, Jiri Damborsky, Michal Kuty, Ivana Kuta Smatanova
| Format: | Article |
|---|---|
| Diterbitkan: | MDPI AG 2019-07-01 |
Deskripsi
Haloalkane dehalogenases are a very important class of microbial enzymes for environmental detoxification of halogenated pollutants, for biocatalysis, biosensing and molecular tagging. The double mutant (Ile44Leu + Gln102His) of the haloalkane dehalogenase DbeA from <i>Bradyrhizobium elkanii</i> USDA94 (DbeAΔCl) was constructed to study the role of the second halide-binding site previously discovered in the wild-type structure. The variant is less active, less stable in the presence of chloride ions and exhibits significantly altered substrate specificity when compared with the DbeAwt. DbeAΔCl was crystallized using the sitting-drop vapour-diffusion procedure with further optimization by the random microseeding technique. The crystal structure of the DbeAΔCl has been determined and refined to the 1.4 Å resolution. The DbeAΔCl crystals belong to monoclinic space group <i>C</i>121. The DbeAΔCl molecular structure was characterized and compared with five known haloalkane dehalogenases selected from the Protein Data Bank.