title: An S188V mutation alters substrate specificity of non-stereospecific α-haloalkanoic acid dehalogenase E (DehE).
abstract: The non-stereospecific α-haloalkanoic acid dehalogenase E (DehE) degrades many halogenated compounds but is ineffective against β-halogenated compounds such as 3-chloropropionic acid (3CP). Using molecular dynamics (MD) simulations and site-directed mutagenesis we show here that introducing the mutation S188V into DehE improves substrate specificity towards 3CP. MD simulations showed that residues W34, F37, and S188 of DehE were crucial for substrate binding. DehE showed strong binding ability for D-2-chloropropionic acid (D-2CP) and L-2-chloropropionic acid (L-2CP) but less affinity for 3CP. This reduced affinity was attributed to weak hydrogen bonding between 3CP and residue S188, as the carboxylate of 3CP forms rapidly interconverting hydrogen bonds with the backbone amide and side chain hydroxyl group of S188. By replacing S188 with a valine residue, we reduced the inter-molecular distance and stabilised bonding of the carboxylate of 3CP to hydrogens of the substrate-binding residues. Therefore, the S188V can act on 3CP, although its affinity is less strong than for D-2CP and L-2CP as assessed by Km. This successful alteration of DehE substrate specificity may promote the application of protein engineering strategies to other dehalogenases, thereby generating valuable tools for future bioremediation technologies.
authors: Department of Biotechnology, Faculty of Science, IIUM, Bandar Indera Mahkota, 25200, Kuantan, Pahang, Malaysia.
authors: Azzmer Azzar Abdul
authors: AA
authors: Hamid
authors: Department of Biotechnology, Faculty of Science, IIUM, Bandar Indera Mahkota, 25200, Kuantan, Pahang, Malaysia.
authors: Tengku Haziyamin Tengku Abdul
authors: TH
authors: Hamid
authors: Department of Chemistry, Faculty of Science, Universiti Teknologi Malaysia, 81310, UTM Johor Bahru, Johor, Malaysia.
authors: Roswanira Abdul
authors: RA
authors: Wahab
authors: Department of Biosciences and Health Sciences, Faculty of Biosciences and Medical Engineering, Universiti Teknologi Malaysia, 81310, UTM Johor Bahru, Johor, Malaysia.
authors: Mohd Shahir Shamsir
authors: MS
authors: Omar
authors: Department of Biotechnology and Medical Engineering, Faculty of Biosciences and Medical Engineering, Universiti Teknologi Malaysia, 81310, UTM Johor Bahru, Johor, Malaysia.
authors: Fahrul
authors: F
authors: Huyop
mesh_terms: D001665:Binding Sites; D006860:Hydrogen Bonding; D006867:Hydrolases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D011422:Propionates / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013379:Substrate Specificity; D014633:Valine / Q000378:metabolism
chemical_list: D011422:Propionates; D012694:Serine; D006867:Hydrolases; C034507:2-haloacid dehalogenase; D014633:Valine; C007022:3-chloropropionic acid