Exploring the Structure and Activity of Haloalkane Dehalogenase from Sphingomonas paucimobilis UT26: Evidence for Product- and Water-Mediated Inhibition
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- 作者：Aaron J. Oakley ; Zbyn
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- 刊名：Biochemistry
- 出版年：2002
- 出版时间：April 16, 2002
- 年：2002
- 卷：41
- 期：15
- 页码：4847 - 4855
- 全文大小：404K
- 年卷期：v.41,no.15(April 16, 2002)
- ISSN：1520-4995
文摘
The hydrolysis of haloalkanes to their corresponding alcohols and inorganic halides is catalyzedby 
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-hydrolases called haloalkane dehalogenases. The study of haloalkane dehalogenases is vital forthe development of these enzymes if they are to be utilized for bioremediation of organohalide-contaminatedindustrial waste. We report the kinetic and structural analysis of the haloalkane dehalogenase fromSphingomonas 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 weakbut 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, forexample, to the activity of LinB with 1-chlorobutane [68.2 nmol s-1 (mg of enzyme)-1]. Inhibition analysisreveals that both 1,2-dichloroethane and 1,2-dichloropropane act as simple competitive inhibitors of thesubstrate 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 andhalide ions confirm that these compounds could bind productively. However, when these moieties wereincluded 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 showthe importance of active site water molecules and reaction products in molecular docking.
/-hydrolases called haloalkane dehalogenases. The study of haloalkane dehalogenases is vital forthe development of these enzymes if they are to be utilized for bioremediation of organohalide-contaminatedindustrial waste. We report the kinetic and structural analysis of the haloalkane dehalogenase fromSphingomonas 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 weakbut 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, forexample, to the activity of LinB with 1-chlorobutane [68.2 nmol s-1 (mg of enzyme)-1]. Inhibition analysisreveals that both 1,2-dichloroethane and 1,2-dichloropropane act as simple competitive inhibitors of thesubstrate 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 andhalide ions confirm that these compounds could bind productively. However, when these moieties wereincluded 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 showthe importance of active site water molecules and reaction products in molecular docking.