Haloalkane dehalogenase (DhlA) converts 1,2-dichloroethane (1,2-DCA) to 2-chloroethane in the genus Ancylobacter and Xanthobacter autotrophicus GJ10 (XaDhlA) and allows these organisms to utilise 1,2-DCA and some other halogenated alkanes for growth. The DhlA encoding gene (dhlA) was PCR-amplified from the genomic DNA of a recently isolated Ancylobacter aquaticus UV5 strain, cloned and overexpressed in Escherichia coli BL21 (DE3). The recombinant enzyme was purified by using Amicon ultra-15 centrifugal filter units, an anion-exchange QFF column followed by a gel-filtration column (Sephacryl HR100). Enzyme activity was determined by using 1,2-DCA as a substrate. Three-dimensional structure of the enzyme was predicted using SWISS-MODEL workspace and the biophysical properties were predicted by submitting the amino acid sequence of DhlA on ExPASy server. DhlA (M-r 35 kDa) exhibited optimum activity at temperature 37 degrees C and pH 9.0. The enzyme retained approximately 50% of its activity after 1 h of incubation at 50 degrees C, and showed moderate stability against denaturing agent urea. The DhlA displayed a K-m value of 842 mu M and k(cat)/K-m ratio of 168 mM(-1) min(-1) for its substrate 1,2-DCA. This DhlA was found to belong to the alpha/beta hydrolase family with a catalytic triad composed of Asp-His-Asp in its active site. This is the first study reporting on the characterisation and reaction kinetics of purified DhlA from A. aquaticus UV5 indigenous to contaminated site in Africa. (C) 2014 Elsevier Inc. All rights reserved.