Structural highlights
Function
[DHLA_XANAU] Catalyzes hydrolytic cleavage of carbon-halogen bonds in halogenated aliphatic compounds, leading to the formation of the corresponding primary alcohols, halide ions and protons. Has a broad substrate specificity, which includes terminally mono- and di- chlorinated and brominated alkanes (up to C4 only). The highest activity was found with 1,2-dichloroethane, 1,3-dichloropropane, and 1,2-dibromoethane.
Evolutionary Conservation
Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf.
Publication Abstract from PubMed
Conversion of halogenated aliphatics by haloalkane dehalogenase proceeds via the formation of a covalent alkyl-enzyme intermediate which is subsequently hydrolyzed by water. In the wild type enzyme, the slowest step for both 1,2-dichloroethane and 1,2-dibromoethane conversion is a unimolecular enzyme isomerization preceding rapid halide dissociation. Phenylalanine 172 is located in a helix-loop-helix structure that covers the active site cavity of the enzyme, interacts with the C1 beta of 1,2-dichloroethane during catalysis, and could be involved in stabilization of this helix-loop-helix region of the cap domain of the enzyme. To obtain more information about the role of this residue in dehalogenase function, we performed a mutational analysis of position 172 and studied the kinetics and X-ray structure of the Phe172Trp enzyme. The Phe172Trp mutant had a 10-fold higher Kcat/Km for 1-chlorohexane and a 2-fold higher Kcat for 1,2-dibromoethane than the wild-type enzyme. The X-ray structure of the Phe172Trp enzyme showed a local conformational change in the helix-loop-helix region that covers the active site. This could explain the elevated activity for 1-chlorohexane of the Phe172Trp enzyme, since it allows this large substrate to bind more easily in the active site cavity. Pre-steady-state kinetic analysis showed that the increase in Kcat found for 1,2-dibromoethane conversion could be attributed to an increase in the rate of an enzyme isomerization step that preceeds halide release. The observed conformational difference between the helix-loop-helix structures of the wild-type enzyme and the faster mutant suggests that the isomerization required for halide release could be a conformational change that takes place in this region of the cap domain of the dehalogenase. It is proposed that Phe172 is involved in stabilization of the helix-loop-helix structure that covers the active site of the enzyme and creates a rigid hydrophobic cavity for small apolar halogenated alkanes.
Kinetic characterization and X-ray structure of a mutant of haloalkane dehalogenase with higher catalytic activity and modified substrate range.,Schanstra JP, Ridder IS, Heimeriks GJ, Rink R, Poelarends GJ, Kalk KH, Dijkstra BW, Janssen DB Biochemistry. 1996 Oct 8;35(40):13186-95. PMID:8855957[1]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Schanstra JP, Ridder IS, Heimeriks GJ, Rink R, Poelarends GJ, Kalk KH, Dijkstra BW, Janssen DB. Kinetic characterization and X-ray structure of a mutant of haloalkane dehalogenase with higher catalytic activity and modified substrate range. Biochemistry. 1996 Oct 8;35(40):13186-95. PMID:8855957 doi:10.1021/bi961151a
