title: Probing the role of amino acids in oxime-mediated reactivation of nerve agent-inhibited human acetylcholinesterase.
abstract: In this study, we employed site-directed mutagenesis to understand the role of amino acids in the gorge in oxime-induced reactivation of nerve agent-inhibited human (Hu) acetylcholinesterase (AChE). The organophosphorus (OP) nerve agents studied included GA (tabun), GB (sarin), GF (cyclosarin), VX, and VR. The kinetics of reactivation were examined using both the mono-pyridinium oxime 2-PAM and bis-pyridinium oximes MMB4, HI-6, and HLö-7. The second-order reactivation rate constants were used to compare reactivation of nerve agent-inhibited wild-type (WT) and mutant enzymes. Residues including Y72, Y124 and W286 were found to play important roles in reactivation by bis-pyridinium, but not by mono-pyridinium oximes. Residue Y124 also was found to play a key role in reactivation by HI-6 and HLö-7, while E202 was important for reactivation by all oximes. Residue substitutions of F295 by Leu and Y337 by Ala showed enhanced reactivation by bis-pyridinium oximes MMB4, HI-6, and HLö-7, possibly by providing more accessibility of the OP moiety associated at the active-site serine to the oxime. These results are similar to those observed previously with bovine AChE and demonstrate that there is significant similarity between human and bovine AChEs with regard to oxime reactivation.
authors.affiliation: Walter Reed Army Institute of Research, Division of Biochemistry, 503 Robert Grant Avenue, Silver Spring, MD 20910-7500, USA; US Military HIV Research Program, Walter Reed Army Institute of Research, Silver Spring, MD 20910-7500, USA.
authors.forename: Carolyn
authors.initials: C
authors.lastname: Chambers
authors.affiliation: Walter Reed Army Institute of Research, Division of Biochemistry, 503 Robert Grant Avenue, Silver Spring, MD 20910-7500, USA.
authors.forename: Chunyuan
authors.initials: C
authors.lastname: Luo
authors.affiliation: Walter Reed Army Institute of Research, Division of Biochemistry, 503 Robert Grant Avenue, Silver Spring, MD 20910-7500, USA.
authors.forename: Min
authors.initials: M
authors.lastname: Tong
authors.affiliation: Walter Reed Army Institute of Research, Division of Biochemistry, 503 Robert Grant Avenue, Silver Spring, MD 20910-7500, USA.
authors.forename: Yerie
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Walter Reed Army Institute of Research, Division of Biochemistry, 503 Robert Grant Avenue, Silver Spring, MD 20910-7500, USA; US Military HIV Research Program, Walter Reed Army Institute of Research, Silver Spring, MD 20910-7500, USA. Electronic address: ashima.saxena@us.army.mil.
authors.forename: Ashima
authors.initials: A
authors.lastname: Saxena
mesh_terms: D000110:Acetylcholinesterase / Q000235:genetics* / Q000378:metabolism; D000596:Amino Acids / Q000235:genetics*; D000818:Animals; D016466:CHO Cells; D002417:Cattle; D002619:Chemical Warfare Agents / Q000633:toxicity*; D002801:Cholinesterase Reactivators / Q000494:pharmacology*; D003412:Cricetulus; D006801:Humans; D009943:Organophosphorus Compounds / Q000633:toxicity*; D010091:Oximes / Q000494:pharmacology; D011220:Pralidoxime Compounds / Q000494:pharmacology; D011726:Pyridinium Compounds / Q000494:pharmacology
chemical_list: D000596:Amino Acids; D002619:Chemical Warfare Agents; D002801:Cholinesterase Reactivators; D009943:Organophosphorus Compounds; D010091:Oximes; D011220:Pralidoxime Compounds; D011726:Pyridinium Compounds; C058577:HLo 7; C005514:N,N'-monomethylenebis(pyridiniumaldoxime); D000110:Acetylcholinesterase; C022870:asoxime chloride; C028797:pralidoxime
keywords: Human AChE mutants; In vitro study; Nerve agents; Oximes; Reactivation; Site-directed mutagenesis