Catalytic antibodies with acetylcholinesterase activity.
We describe three catalytic cholinesterase-like catalytic antibodies (Ab1), as well as anti-idiotypic (Ab2) and idiotypic (Ab3) antibodies, to one of the Ab1s. The Ab1s were raised against the human erythrocyte acetylcholinesterase (AChE), and are unusual in that they both recognise and resemble acetylcholinesterase in their catalytic activity. No contamination of the antibody preparations with either acetylcholinesterase or butyrylcholinesterase (BChE) was found. None of the Ab2s showed catalytic activity, whereas four Ab3s did (an incidence of 1.26% of all Ab3s). Although there is considerable resemblance between Ab1s and Ab3s, there are significant differences between the two groups. All the antibodies were inhibited by phenylmethylsulphonyl fluoride (PMSF), indicating the presence of a serine residue in their active sites, and were inhibited by the cholinesterase active site inhibitors iso-OMPA and pyridostigmine, suggesting the similarity of the sites to those of cholinesterases. The Ab3s resemble the Ab1s in their ability to hydrolyse both acetyl and butyrylthiocholine (BTCh). However, the Ab3s appear to be better catalysts, having significantly reduced K(m) values (for acetyl, but not for butyrylthiocholine) and increased turnover numbers (K(cat)), rate enhancements (K(cat)/K(uncat)) and K(cat)/K(m) ratios, for both substrates, although these values by no means approach those of the natural enzymes. The Ab1s appear to have structures resembling the anionic sites of cholinesterases, as shown by their reaction with the anionic site inhibitors (edrophonium and tetramethylammonium). No such reactions were observed in the Ab3s. None of the antibodies show evidence of the sites resembling the peripheral anionic site (PAS) of acetylcholinesterase. All the antibodies recognise, to varying degrees, the peripheral anionic site of acetylcholinesterase. This was shown by their ability to inhibit acetylcholinesterase, to compete with peripheral site inhibitors, and to block acetylcholinesterase-mediated cell adhesion, a property of this site. The results indicate idiotypic mimicry of a catalytic antibody's active site, and suggest that the development of the catalytic activity in the anti-acetylcholinesterase antibodies may be related to the structural features of the peripheral anionic site of acetylcholinesterase.
Johnson
Glynis
G
Department of Pediatric Surgery/Medical Biochemistry, University of Stellenbosch, P.O. Box 19063, Tygerberg 7505, South Africa. gjo@maties.sun.ac.za
Moore
Samuel W
SW
D000109:Acetylcholine / Q000378:metabolism; D000110:Acetylcholinesterase / Q000276:immunology*; D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000378:metabolism*; D017151:Antibodies, Catalytic / Q000378:metabolism*; D001667:Binding, Competitive; D002092:Butyrylthiocholine / Q000378:metabolism; D002448:Cell Adhesion; D002460:Cell Line; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007130:Immunoglobulin Idiotypes / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D008807:Mice, Inbred BALB C
D000888:Antibodies, Anti-Idiotypic; D017151:Antibodies, Catalytic; D007130:Immunoglobulin Idiotypes; D002092:Butyrylthiocholine; D000110:Acetylcholinesterase; D000109:Acetylcholine