title: Partial characterization of an enzyme that hydrolyzes sarin, soman, tabun, and diisopropyl phosphorofluoridate (DFP).
abstract: The properties of a rat liver enzyme that hydrolyzes organophosphorus (OP) inhibitors of cholinesterases were studied. The rates of hydrolysis of OP inhibitors were determined by continuous titration of released hydrogen ions, using a pH stat method. Centrifugation of homogenates at 205,000 g for 30 min demonstrated that the activity was in the soluble fraction. Hydrolysis of sarin, soman, and diisopropyl phosphorofluoridate (DFP), but not of tabun, was stimulated by the addition of Mn2+ and Mg2+. Hydrolysis of sarin greater than soman greater than tabun greater than DFP. Unlike other OP hydrolases that preferentially hydrolyze the non-toxic isomers of soman, this enzyme hydrolyzed all four soman isomers at approximately the same rate. This result was obtained in vitro by gas chromatographic analysis of enzyme-catalyzed soman hydrolysis and confirmed in vivo by demonstrating reduced toxicity in mice of soman partially hydrolyzed by this enzyme. Km and Vmax were determined by fitting V vs [S] to a hyperbolic function using regression analysis. Km values ranged from 1.1 mM for soman to 8.9 mM for tabun. Vmax values ranged from 54 nmol/min/mg protein for DFP to 2694 for sarin. The enzyme was stable for at least 2 months at -90 degrees but was inactivated by heating at 100 degrees for 5 min. Elution profiles from gel filtration by high pressure liquid chromatography showed that the hydrolytic activity for the OP inhibitors eluted in a single peak, suggesting that a single enzyme was responsible for the observed hydrolysis. Further purification and characterization of this enzyme should prove useful for the development of methods for detection, detoxification, and decontamination of these cholinesterase inhibitors.
authors.affiliation: United States Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, MD 21010-5425.
authors.forename: J S
authors.initials: JS
authors.lastname: Little
authors.forename: C A
authors.initials: CA
authors.lastname: Broomfield
authors.forename: M K
authors.initials: MK
authors.lastname: Fox-Talbot
authors.forename: L J
authors.initials: LJ
authors.lastname: Boucher
authors.forename: B
authors.initials: B
authors.lastname: MacIver
authors.forename: D E
authors.initials: DE
authors.lastname: Lenz
mesh_terms: D000818:Animals; D006867:Hydrolases / Q000032:analysis*; D006868:Hydrolysis; D007531:Isoflurophate / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D010755:Organophosphates / Q000378:metabolism*; D009943:Organophosphorus Compounds / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D012524:Sarin / Q000378:metabolism*; D012999:Soman / Q000378:metabolism*; D013237:Stereoisomerism
chemical_list: D010755:Organophosphates; D009943:Organophosphorus Compounds; D007531:Isoflurophate; D012999:Soman; D012524:Sarin; D006867:Hydrolases; D008274:Magnesium; C009374:tabun