Purification and properties of the membrane-bound form of acetylcholinesterase from chicken brain. Evidence for two distinct polypeptide chains.
The major molecular form of acetylcholinesterase (AChE) from chicken brain is a membrane-bound glycoprotein with an apparent sedimentation coefficient of 11.4 S. Analysis of the purified protein by gel filtration, velocity sedimentation, and sodium dodecyl sulfate-gel electrophoresis shows that the solubilized enzyme is a globular tetramer with an apparent Mr = 420,000. This membrane-bound form of AChE is hydrophobic and readily aggregates in the absence of detergent. These aggregates are concentration-dependent, relatively stable in the presence of high salt concentrations, yet readily dissociate upon addition of detergent to the 11.4 S form, indicating that the interactions are hydrophobic. Polyclonal and monoclonal antibodies raised against chicken brain AChE purified by ion exchange chromatography, affinity chromatography, and preparative gel electrophoresis precipitate AChE enzyme activity. However, these antibodies do not cross-react with the enzyme from chicken muscle which preferentially hydrolyses butyrylcholine. Immunoprecipitation of isotopically labeled enzyme molecules from tissue cultured brain cells and analysis by sodium dodecyl sulfate-gel electrophoresis shows that AChE consists of two polypeptide chains with apparent Mr = 105,000 (alpha) and 100,000 (beta) in a 1:1 ratio. Immunoblotting of brain AChE with either the polyclonal or monoclonal antibodies indicates that the alpha and beta chains share antigenic determinants. Furthermore, both polypeptide chains can be labeled with [3H]diisopropyl fluorophosphate, indicating that they each contain a catalytic site. This is the first indication that globular forms of AChE may consist of multiple polypeptide chains.
D000110:Acetylcholinesterase / Q000302:isolation & purification* / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002462:Cell Membrane / Q000201:enzymology; D002645:Chickens; D003902:Detergents / Q000494:pharmacology; D004789:Enzyme Activation; D000939:Epitopes / Q000032:analysis; D007106:Immune Sera; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D013045:Species Specificity
D003902:Detergents; D000939:Epitopes; D007106:Immune Sera; D046911:Macromolecular Substances; D011092:Polyethylene Glycols; D017830:Octoxynol; D000110:Acetylcholinesterase