Chromatographic characterization of neurotoxic esterase

Neurotoxic esterase (neuropathy target enzyme, NTE) is an enzyme whose irreversible inhibition is the apparent first step in the induction of organophosphorus-induced delayed neuropathy. NTE is an integral membrane protein and thus must be solubilized before isolation can be attempted. This study describes solubilization of active chicken brain NTE with the nondenaturing detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) and characterization of the detergent-solubilized enzyme by gel exclusion chromatography. When detergent-solubilized membranes were chromatographed on Sepharose gel exclusion media, NTE activity eluted with an apparent molecular weight of 880-970 kD. When [3H]diisopropylphosphorofluoridate-radiolabeled membranes and unlabeled microsomal membranes were CHAPS-solubilized, combined and chromatographed on Sepharose 4B, NTE activity coeluted with two radiolabeled proteins (Mr = 148 kD and Mr = 112 kD using sodium dodecyl sulfate-polyacrylamide gel electrophoresis with reducing conditions). Another radiolabeled protein (Mr = 92 kD) coeluted exclusively with inhibitor-resistant esterase activity. This study provides strong evidence that the 148 and 112 kD proteins are subunits of a multicomponent NTE complex.