title: Purification of soluble acetylcholinesterase from sheep liver by affinity chromatography.
abstract: The purpose of this study was to develop a protocol for the purification of acetylcholinesterase (AChE, acetylcholine acetylhydrolase, E.C.3.1.1.7) enzyme and to extend a purification method for further enzyme characterization. A further aim was to study whether the edrophonium's pharmacologic action is due primarily to the inhibition or inactivation of AChE at sites of cholinergic transmission. The purification of a soluble AChE from sheep liver using affinity chromatography on Concanavalin A-Sepharose 4B and edrophonium-Sepharose 6B is studied. The affinity matrix was synthesized by coupling an inhibitor edrophonium to epoxy-activated Sepharose at flow rate of 0.5 ml/min. AChE is a pivotal enzyme in the cholinergic nervous system. Its primary function is to catalyze hydrolysis of released acetylcholine (ACh) and thus maintain homeostasis of this neurotransmitter in the central and peripheral nervous systems. Hence, AChE is important in both pharmacological and toxicological mechanisms. It was purified 842-fold with a specific activity of 21 U/mg protein. Sodium dodecyl sulfate (SDS) electrophoresis resulted in a monomeric molecular weight of 67.04 kDa, while on gel chromatography using Sephacryl S-200 under nondenaturing conditions to be 201.5 kDa. Based on the molecular weight obtained by gel filtration, the purified AChE was assumed to be a tetrameric form.
authors.affiliation: School of Biomedical and Biological Sciences, Faculty of Science and Technology, University of Plymouth, Plymouth, PL4 8AA, UK. kasim.abassaskar@plymouth.ac.uk
authors.forename: Kasim Abass
authors.initials: KA
authors.lastname: Askar
authors.forename: A Caleb
authors.initials: AC
authors.lastname: Kudi
authors.forename: A John
authors.initials: AJ
authors.lastname: Moody
mesh_terms: D000110:Acetylcholinesterase / Q000737:chemistry* / Q000302:isolation & purification*; D000818:Animals; D002846:Chromatography, Affinity / Q000379:methods*; D002850:Chromatography, Gel; D012756:Sheep; D012995:Solubility
chemical_list: D000110:Acetylcholinesterase