Allosteric control of acetylcholinesterase activity by monoclonal antibodies.
Previous studies showed that monoclonal antibodies raised against phosphorylated fetal bovine serum acetylcholinesterase appeared to modulate the catalytic activity of the enzyme by binding to a conformational epitope located at or near the region of the peripheral anionic site. The mechanism of inhibition of acetylcholinesterase by these monoclonal antibodies was further investigated by determining their effect on (i) substrate inhibition due to the binding of excess substrate to the peripheral anionic site and (ii) binding of peripheral anionic site ligands, such as propidium and fasciculin. Results of these experiments demonstrate that the accessibility of substrate to the peripheral anionic site in these complexes was restricted but not completely blocked, as none of the monoclonal antibodies eliminated the phenomenon of excess substrate inhibition. The results also show that propidium clearly slowed the inhibition of fetal bovine serum acetylcholinesterase by all six inhibitory monoclonal antibodies but to different levels. Complexation of fetal bovine serum acetylcholinesterase with monoclonal antibodies 25B1, 4E5, 6H9, and 5E8 interfered with the binding of fasciculin to the complexed enzyme, suggesting that part of their epitope overlapped with the fasciculin binding site. These monoclonal antibodies bind, in part, at the peripheral anionic site, since polyclonal anti-idiotypic antibodies generated against two monoclonal antibodies, 25B1 and 6H9, bound stoichiometric amounts of propidium. Like fasciculin, binding of these monoclonal antibodies in the vicinity of the peripheral anionic site at the rim of the active site gorge allosterically affects the orientation of W86 located at the base of the gorge, resulting in inhibition of enzyme activity.
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Division of Biochemistry, Walter Reed Army Institute of Research, Washington, DC 20307-5100, USA.
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D000110:Acetylcholinesterase / Q000276:immunology* / Q000378:metabolism*; D000494:Allosteric Regulation / Q000276:immunology; D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000737:chemistry; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology*; D000915:Antibody Affinity; D001666:Binding Sites, Antibody; D001667:Binding, Competitive / Q000276:immunology; D002417:Cattle; D002800:Cholinesterase Inhibitors / Q000276:immunology / Q000378:metabolism; D004546:Elapid Venoms / Q000276:immunology / Q000378:metabolism; D004789:Enzyme Activation / Q000276:immunology; D005312:Fetal Blood / Q000201:enzymology / Q000276:immunology; D046911:Macromolecular Substances; D011485:Protein Binding / Q000276:immunology; D011817:Rabbits; D013379:Substrate Specificity
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D002800:Cholinesterase Inhibitors; D004546:Elapid Venoms; D046911:Macromolecular Substances; C044937:fasciculin; D000110:Acetylcholinesterase