Exploring the structure and activity of haloalkane dehalogenase from Sphingomonas paucimobilis UT26: Evidence for product- and water-mediated inhibition

Aaron J. Oakley, Zbyněk Prokop, Michal Boháč, Jan Kmuníček, Tomáš Jedlička, Marta Monincová, Ivana Kutá-Smatanová, Yuji Nagata, Jiří Damborský, Matthew C.J. Wilce

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The hydrolysis of haloalkanes to their corresponding alcohols and inorganic halides is catalyzed by α/β-hydrolases called haloalkane dehalogenases. The study of haloalkane dehalogenases is vital for the development of these enzymes if they are to be utilized for bioremediation of organohalide-contaminated industrial waste. We report the kinetic and structural analysis of the haloalkane dehalogenase from Sphingomonas paucimobilis UT26 (LinB) in complex with each of 1,2-dichloroethane and 1,2-dichloropropane and the reaction product of 1-chlorobutane turnover. Activity studies showed very weak but detectable activity of LinB with 1,2-dichloroethane [0.012 nmol s-1 (mg of enzyme)-1] and 1,2-dichloropropane [0.027 nmol s-1 (mg of enzyme)-1]. These activities are much weaker compared, for example, to the activity of LinB with 1-chlorobutane [68.2 nmol s-1 (mg of enzyme)-1]. Inhibition analysis reveals that both 1,2-dichloroethane and 1,2-dichloropropane act as simple competitive inhibitors of the substrate 1-chlorobutane and that 1,2-dichloroethane binds to LinB with lower affinity than 1,2- dichloropropane. Docking calculations on the enzyme in the absence of active site water molecules and halide ions confirm that these compounds could bind productively. However, when these moieties were included in the calculations, they bound in a manner similar to that observed in the crystal structure. These data provide an explanation for the low activity of LinB with small, chlorinated alkanes and show the importance of active site water molecules and reaction products in molecular docking.

Original languageEnglish
Pages (from-to)4847-4855
Number of pages9
Issue number15
Publication statusPublished - 16 Apr 2002
Externally publishedYes

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