title: Direct detection of stereospecific soman hydrolysis by wild-type human serum paraoxonase.
abstract: Human serum paraoxonase 1 (HuPON1; EC 3.1.8.1) is a calcium-dependent six-fold beta-propeller enzyme that has been shown to hydrolyze an array of substrates, including organophosphorus (OP) chemical warfare nerve agents. Although recent efforts utilizing site-directed mutagenesis have demonstrated specific residues (such as Phe222 and His115) to be important in determining the specificity of OP substrate binding and hydrolysis, little effort has focused on the substrate stereospecificity of the enzyme; different stereoisomers of OPs can differ in their toxicity by several orders of magnitude. For example, the C+/-P- isomers of the chemical warfare agent soman (GD) are known to be more toxic by three orders of magnitude. In this study, the catalytic activity of HuPON1 towards each of the four chiral isomers of GD was measured simultaneously via chiral GC/MS. The catalytic efficiency (k(cat)/K(m)) of the wild-type enzyme for the various stereoisomers was determined by a simultaneous solution of hydrolysis kinetics for each isomer. Derived k(cat)/K(m) values ranged from 625 to 4130 mm(-1).min(-1), with isomers being hydrolyzed in the order of preference C+P+ > C-P+ > C+P- > C-P-. The results indicate that HuPON1 hydrolysis of GD is stereoselective; substrate stereospecificity should be considered in future efforts to enhance the OPase activity of this and other candidate bioscavenger enzymes.
authors.affiliation: Physiology and Immunology Branch, Research Division, US Army Medical Research Institute of Chemical Defense, 3100 Ricketts Point Road, Aberdeen Proving Ground, MD 21010-5400, USA.
authors.forename: David T
authors.initials: DT
authors.lastname: Yeung
authors.forename: J Richard
authors.initials: JR
authors.lastname: Smith
authors.forename: Richard E
authors.initials: RE
authors.lastname: Sweeney
authors.forename: David E
authors.initials: DE
authors.lastname: Lenz
authors.forename: Douglas M
authors.initials: DM
authors.lastname: Cerasoli
mesh_terms: D043303:Aryldialkylphosphatase / Q000097:blood* / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D002384:Catalysis; D002619:Chemical Warfare Agents / Q000032:analysis / Q000737:chemistry / Q000378:metabolism* / Q000633:toxicity; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D012999:Soman / Q000032:analysis / Q000737:chemistry / Q000378:metabolism* / Q000633:toxicity; D013237:Stereoisomerism; D013379:Substrate Specificity
chemical_list: D002619:Chemical Warfare Agents; D012999:Soman; D043303:Aryldialkylphosphatase; C468406:PON1 protein, human