Family Report for: Haloacetate_dehalogenase

As in epoxide hydrolase and haloalkane dehalogenase the catalytic triad nucleophile is an invariable aspartate. The region around the nucleophile aspartate is GHDRG. See van Loo 2006 Appl.Environ.Microbiol 72 2905

Title: X-Ray crystallographic and mutational studies of fluoroacetate dehalogenase from Burkholderia sp. strain FA1
Jitsumori K, Omi R, Kurihara T, Kurata A, Mihara H, Miyahara I, Hirotsu K, Esaki N
Ref: Journal of Bacteriology, 191:2630, 2009 : PubMed
Abstract


ESTHER: Jitsumori_2009_J.Bacteriol_191_2630
PubMedSearch: Jitsumori 2009 J.Bacteriol 191 2630
PubMedID: 19218394
Gene_locus related to this paper: bursp-deha

Abstract

Fluoroacetate dehalogenase catalyzes the hydrolytic defluorination of fluoroacetate to produce glycolate. The enzyme is unique in that it catalyzes the cleavage of a carbon-fluorine bond of an aliphatic compound: the bond energy of the carbon-fluorine bond is among the highest found in natural products. The enzyme also acts on chloroacetate, although much less efficiently. We here determined the X-ray crystal structure of the enzyme from Burkholderia sp. strain FA1 as the first experimentally determined three-dimensional structure of fluoroacetate dehalogenase. The enzyme belongs to the alpha/beta hydrolase superfamily and exists as a homodimer. Each subunit consists of core and cap domains. The catalytic triad, Asp104-His271-Asp128, of which Asp104 serves as the catalytic nucleophile, was found in the core domain at the domain interface. The active site was composed of Phe34, Asp104, Arg105, Arg108, Asp128, His271, and Phe272 of the core domain and Tyr147, His149, Trp150, and Tyr212 of the cap domain. An electron density peak corresponding to a chloride ion was found in the vicinity of the N(epsilon1) atom of Trp150 and the N(epsilon2) atom of His149, suggesting that these are the halide ion acceptors. Site-directed replacement of each of the active-site residues, except for Trp150, by Ala caused the total loss of the activity toward fluoroacetate and chloroacetate, whereas the replacement of Trp150 caused the loss of the activity only toward fluoroacetate. An interaction between Trp150 and the fluorine atom is probably an absolute requirement for the reduction of the activation energy for the cleavage of the carbon-fluorine bond.



        

Title: Diversity and biocatalytic potential of epoxide hydrolases identified by genome analysis
van Loo B, Kingma J, Arand M, Wubbolts MG, Janssen DB
Ref: Applied Environmental Microbiology, 72:2905, 2006 : PubMed
Abstract


ESTHER: van Loo_2006_Appl.Environ.Microbiol_72_2905
PubMedSearch: van Loo 2006 Appl.Environ.Microbiol 72 2905
PubMedID: 16597997
Gene_locus related to this paper: agrra-echA, bacsu-yfhM, corsp-cEH, deira-DR2549, myctu-ephF, nosp7-b2j4w6, nosp7-b2iv02, psepu-PP4624, rhopa-q6n645, strco-2SCG4.13, thefy-q47qj2

Abstract

Epoxide hydrolases play an important role in the biodegradation of organic compounds and are potentially useful in enantioselective biocatalysis. An analysis of various genomic databases revealed that about 20% of sequenced organisms contain one or more putative epoxide hydrolase genes. They were found in all domains of life, and many fungi and actinobacteria contain several putative epoxide hydrolase-encoding genes. Multiple sequence alignments of epoxide hydrolases with other known and putative alpha/beta-hydrolase fold enzymes that possess a nucleophilic aspartate revealed that these enzymes can be classified into eight phylogenetic groups that all contain putative epoxide hydrolases. To determine their catalytic activities, 10 putative bacterial epoxide hydrolase genes and 2 known bacterial epoxide hydrolase genes were cloned and overexpressed in Escherichia coli. The production of active enzyme was strongly improved by fusion to the maltose binding protein (MalE), which prevented inclusion body formation and facilitated protein purification. Eight of the 12 fusion proteins were active toward one or more of the 21 epoxides that were tested, and they converted both terminal and nonterminal epoxides. Four of the new epoxide hydrolases showed an uncommon enantiopreference for meso-epoxides and/or terminal aromatic epoxides, which made them suitable for the production of enantiopure (S,S)-diols and (R)-epoxides. The results show that the expression of epoxide hydrolase genes that are detected by analyses of genomic databases is a useful strategy for obtaining new biocatalysts.