title: Structure of Rhodococcus erythropolis limonene-1,2-epoxide hydrolase reveals a novel active site.
abstract: Epoxide hydrolases are essential for the processing of epoxide-containing compounds in detoxification or metabolism. The classic epoxide hydrolases have an alpha/beta hydrolase fold and act via a two-step reaction mechanism including an enzyme-substrate intermediate. We report here the structure of the limonene-1,2-epoxide hydrolase from Rhodococcus erythropolis, solved using single-wavelength anomalous dispersion from a selenomethionine-substituted protein and refined at 1.2 A resolution. This enzyme represents a completely different structure and a novel one-step mechanism. The fold features a highly curved six-stranded mixed beta-sheet, with four alpha-helices packed onto it to create a deep pocket. Although most residues lining this pocket are hydrophobic, a cluster of polar groups, including an Asp-Arg-Asp triad, interact at its deepest point. Site-directed mutagenesis supports the conclusion that this is the active site. Further, a 1.7 A resolution structure shows the inhibitor valpromide bound at this position, with its polar atoms interacting directly with the residues of the triad. We suggest that several bacterial proteins of currently unknown function will share this structure and, in some cases, catalytic properties.
authors.affiliation: Department of Toxicology, University of Würzburg, Versbacher Strasse 9, D-97078 Würzburg, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Arand
authors.forename: B Martin
authors.initials: BM
authors.lastname: Hallberg
authors.forename: Jinyu
authors.initials: J
authors.lastname: Zou
authors.forename: Terese
authors.initials: T
authors.lastname: Bergfors
authors.forename: Franz
authors.initials: F
authors.lastname: Oesch
authors.forename: Mariët J
authors.initials: MJ
authors.lastname: van der Werf
authors.forename: Jan A M
authors.initials: JA
authors.lastname: de Bont
authors.forename: T Alwyn
authors.initials: TA
authors.lastname: Jones
authors.forename: Sherry L
authors.initials: SL
authors.lastname: Mowbray
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D019281:Dimerization; D004851:Epoxide Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012240:Rhodococcus / Q000201:enzymology* / Q000235:genetics; D017386:Sequence Homology, Amino Acid
chemical_list: D001426:Bacterial Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; D004851:Epoxide Hydrolases; C115926:limonene-1,2-epoxide hydrolase