title: L-2-Haloacid dehalogenase from Ancylobacter aquaticus UV5: Sequence determination and structure prediction.
abstract: A novel 25 kDa L-2-haloacid dehalogenase (L-2-DhlB) from a recently isolated Ancylobacter aquaticus strain UV5 indigenous to contaminated site in South Africa is reported here with its gene sequence. The enzyme was purified to 22.1-fold increase in specific activity of 72.9 U/mg protein when the organism was grown in medium supplemented with 5 mM 1,2-dichloroethane (1,2-DCA). L-2-DhlB was optimally active at pH 9.0 and 37°C with poor stability at 50°C, retaining 50% of its activity after 30 min, but inactivated rapidly at 60°C. L-2-DhlB catalyzed monochloroacetate (MCA) with Km and Vmax values of 0.47 mM and 2.4 μM/min, respectively. L-2-DhlB exhibited the kcat value of 4.8/min. Expression of about 100% relative activity of L-2-DhlB on the substrate L-2-monochloropropionate (L-2-MCPA) as compared to 5% on D-2-monochloropropionate (D-2-MCPA) suggested that L-2-DhlB belongs to the family of L-2-haloacid dehalogenases. ES-mass spectroscopy and bioinformatics tools resulted in 693 bp ORF sequence corresponding to 230 amino acid protein. NCBI-BLAST of L-2-DhlB resulted in the detection of a putative conserved domain of hypothetical haloacid dehalogenase (HAD)-like superfamily and subfamily IA.
authors.affiliation: Discipline of Microbiology, School of Life Sciences, College of Agriculture, Engineering and Science, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban 4000, South Africa.
authors.forename: Ajit
authors.initials: A
authors.lastname: Kumar
authors.affiliation: Discipline of Microbiology, School of Life Sciences, College of Agriculture, Engineering and Science, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban 4000, South Africa.
authors.forename: Balakrishna
authors.initials: B
authors.lastname: Pillay
authors.affiliation: Discipline of Microbiology, School of Life Sciences, College of Agriculture, Engineering and Science, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban 4000, South Africa. Electronic address: olanirana@ukzn.ac.za.
authors.forename: Ademola O
authors.initials: AO
authors.lastname: Olaniran
mesh_terms: D020561:Alphaproteobacteria / Q000201:enzymology* / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D019295:Computational Biology; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D006867:Hydrolases; C034507:2-haloacid dehalogenase
keywords: 1,2-Dichloroethane; Ancylobacter aquaticus strain UV5; l-2-Haloacid dehalogenase