Amphiphilic forms of butyrylcholinesterase in mucosal cells of rat intestine.
The properties of a cholinesterase from mucosal cells of rat intestine have been characterized. The enzyme was identified as butyrylcholinesterase because it was more sensitive to iso-OMPA (IC50 = 1.0 x 10(-6) M) than to BW284C51 (IC50 = 5.5 x 10(-5) M) and was not inhibited by substrate excess. It displayed a higher affinity for acetylthiocholine than for butyrylthiocholine. A major molecular form was observed sedimenting at 5.9 S. Two other minor molecular forms were identified as a hydrophilic tetramer (G4, sedimenting at 10.5 S) and a monomer (G1, sedimenting at 4.3 S). The 5.9 S component was referred to as "G" form (G for globular) and not "G2" as usual dimers for the following reasons: (i) the G form was unaffected by the reducing agents, beta-mercaptoethanol and dithiothreitol, which converted disulfide-linked dimers of acetylcholinesterase into monomers, (ii) the G form was shifted from 5.9 to 3.4 S when the sucrose gradient contained Triton X-100. This value of 3.4 S (in Triton X-100) appeared too low for a typical G2 form. The shift in the S value was partly reversible: the 3.4 S form resedimented at 5.2 S in the absence of detergent. The behavior of the G form in sucrose gradients indicated that it was amphiphilic. This was confirmed in nondenaturing electrophoreses and also by quantitative binding of the G form to octyl-Sepharose. The hydrophobic domain of the G form was not a glycolipid, as shown by its insensitivity to Bacillus thuringiensis phosphatidylinositol-specific phospholipase C and its nonaggregating properties in the absence of nondenaturing detergent.(ABSTRACT TRUNCATED AT 250 WORDS)
Sine
J P
JP
Laboratoire de Biochimie II, Faculté des Sciences, Centre de Recherche de Biologie et Physico-Chimie Cellulaires, Nantes, France.
Toutant
J P
JP
Weigel
P
P
Colas
B
B
D000818:Animals; D001665:Binding Sites; D002091:Butyrylcholinesterase / Q000737:chemistry / Q000378:metabolism*; D002499:Centrifugation, Density Gradient; D002800:Cholinesterase Inhibitors / Q000494:pharmacology; D002845:Chromatography; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism; D007413:Intestinal Mucosa / Q000201:enzymology*; D007700:Kinetics; D046911:Macromolecular Substances; D008297:Male; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012995:Solubility
D002800:Cholinesterase Inhibitors; D046911:Macromolecular Substances; D010446:Peptide Fragments; D002091:Butyrylcholinesterase; D010450:Endopeptidases