title: Exploring the aryl esterase catalysis of paraoxonase-1 through solvent kinetic isotope effects and phosphonate-based isosteric analogues of the tetrahedral reaction intermediate.
abstract: Although a recent study of Debord et al. in Biochimie (2014; 97:72-77) described the thermodynamics of the catalysed hydrolysis of phenyl acetate by human paraoxonase-1, the mechanistic details along the reaction route of this enzyme remain unclear. Therefore, we briefly present the solvent kinetic isotope effects on the phenyl acetate esterase activity of paraoxonase-1 and its inhibition with the phenyl methylphosphonate anion, which is a stable isosteric analogue that mimics the high-energy tetrahedral intermediate on the hydroxide-promoted hydrolysis pathway. The data show normal isotope effects, while proton inventory analysis indicates that two protons contribute to the kinetic isotope effect. Coherently, moderate competitive inhibition with the phenyl methylphosphonate anion reveals that the rate-limiting transition state suboptimally resembles the tetrahedral intermediate. The implications of these findings can be attributed to two possible reaction mechanisms that might occur during the paraoxonase-1-catalysed hydrolysis of phenyl acetate.
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov trg 2, 1000 Ljubljana, Slovenia.
authors.forename: Aljoša
authors.initials: A
authors.lastname: Bavec
authors.affiliation: Chair of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Ljubljana, Aškerčeva 7, 1000 Ljubljana, Slovenia.
authors.forename: Damijan
authors.initials: D
authors.lastname: Knez
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov trg 2, 1000 Ljubljana, Slovenia.
authors.forename: Tomaž
authors.initials: T
authors.lastname: Makovec
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov trg 2, 1000 Ljubljana, Slovenia.
authors.forename: Jure
authors.initials: J
authors.lastname: Stojan
authors.affiliation: Chair of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Ljubljana, Aškerčeva 7, 1000 Ljubljana, Slovenia.
authors.forename: Stanislav
authors.initials: S
authors.lastname: Gobec
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov trg 2, 1000 Ljubljana, Slovenia. Electronic address: marko.golicnik@mf.uni-lj.si.
authors.forename: Marko
authors.initials: M
authors.lastname: Goličnik
mesh_terms: D000085:Acetates / Q000737:chemistry / Q000378:metabolism*; D043303:Aryldialkylphosphatase / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D004950:Esterases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D006878:Hydroxides / Q000737:chemistry / Q000378:metabolism; D007554:Isotopes; D007700:Kinetics; D008954:Models, Biological; D008956:Models, Chemical; D015394:Molecular Structure; D063065:Organophosphonates / Q000737:chemistry / Q000378:metabolism*; D010636:Phenols / Q000737:chemistry / Q000378:metabolism*; D011522:Protons; D011994:Recombinant Proteins / Q000378:metabolism; D012997:Solvents; D013379:Substrate Specificity
chemical_list: D000085:Acetates; D006878:Hydroxides; D007554:Isotopes; D063065:Organophosphonates; D010636:Phenols; D011522:Protons; D011994:Recombinant Proteins; D012997:Solvents; C570634:phenyl acetate; D004950:Esterases; D043303:Aryldialkylphosphatase; C468406:PON1 protein, human
keywords: Enzyme reaction mechanism; Inventory proton analysis; Paraoxonase-1; Solvent kinetic isotope effect