title: Characterization of a phosphotriesterase-like lactonase from the hyperthermoacidophilic crenarchaeon Vulcanisaeta moutnovskia.
abstract: The phosphotriesterase-like lactonase (PLL) encoded by Vmut_2255 in the hyperthermoacidophilic crenarchaeon Vulcanisaeta moutnovskia (VmutPLL), represents the only hyperthermophilic PLL homologue identified so far in addition to the previously characterized thermophilic PLLs from Sulfolobus spp. The Vmut_2255 gene was cloned, heterologously expressed in Escherichia coli; the resultant protein purified and characterized as a 82kDa homodimer (36kDa subunits). The VmutPLL converted lactones and acyl-homoserine lactones (AHLs) with comparable activities. Towards organophosphates (OP) VmutPLL showed a promiscuous but significantly lower activity and only minor activity was observed with carboxylesters. The catalytic activity strictly depended on bivalent cations (Cd(2+)>Ni(2+)>Co(2+)>Mn(2+)>Zn(2+)). Furthermore, VmutPLL showed a pH optimum around 8.0, a temperature optimum of 80°C, and thermostability with a half-life of 26min at 90°C. In this work, the stereoselectivity of a PLL enzyme was investigated for the first time using enantiopure lactones. The VmutPLL showed a slight preference but not an exclusive specificity for the (R)-enantiomers of capro- and valerolactone. The high thermal stability as well as the broad substrate spectrum towards lactones, AHLs and OPs underlines the high biotechnological potential of VmutPLL.
authors.affiliation: Molecular Enzyme Technology and Biochemistry, Biofilm Centre, Faculty of Chemistry, University of Duisburg-Essen, Universitaetsstrasse 5, 45141 Essen, Germany. Electronic address: Verena.Kallnik@uni-due.de.
authors.forename: Verena
authors.initials: V
authors.lastname: Kallnik
authors.affiliation: Sigma-Aldrich, Riedstraße 2, D-89555 Steinheim, Germany.
authors.forename: Alina
authors.initials: A
authors.lastname: Bunescu
authors.affiliation: The Henry Wellcome Building for Biocatalysis, Biosciences, College of Life and Environmental Sciences, University of Exeter, United Kingdom.
authors.forename: Christopher
authors.initials: C
authors.lastname: Sayer
authors.affiliation: Molecular Enzyme Technology and Biochemistry, Biofilm Centre, Faculty of Chemistry, University of Duisburg-Essen, Universitaetsstrasse 5, 45141 Essen, Germany.
authors.forename: Christopher
authors.initials: C
authors.lastname: Bräsen
authors.affiliation: Sigma-Aldrich, Industriestrasse 25, CH-9470 Buchs, Switzerland.
authors.forename: Roland
authors.initials: R
authors.lastname: Wohlgemuth
authors.affiliation: The Henry Wellcome Building for Biocatalysis, Biosciences, College of Life and Environmental Sciences, University of Exeter, United Kingdom.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Littlechild
authors.affiliation: Molecular Enzyme Technology and Biochemistry, Biofilm Centre, Faculty of Chemistry, University of Duisburg-Essen, Universitaetsstrasse 5, 45141 Essen, Germany.
authors.forename: Bettina
authors.initials: B
authors.lastname: Siebers
mesh_terms: D002265:Carboxylic Ester Hydrolases / Q000235:genetics / Q000378:metabolism*; D002413:Cations, Divalent; D003001:Cloning, Molecular; D004795:Enzyme Stability; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007783:Lactones / Q000378:metabolism*; D044345:Phosphoric Triester Hydrolases / Q000378:metabolism*; D013379:Substrate Specificity; D019624:Thermoproteaceae / Q000201:enzymology* / Q000235:genetics
chemical_list: D002413:Cations, Divalent; D007783:Lactones; D002265:Carboxylic Ester Hydrolases; D044345:Phosphoric Triester Hydrolases
keywords: Acyl-homoserine lactone; Archaea; Organophosphate detoxification; Thermostable phosphotriesterase-like lactonases; Vulcanisaeta moutnovskia