Neurotoxic esterase: characterization of the solubilized enzyme and the conditions for its solubilization from chicken brain microsomal membranes with ionic, zwitterionic, or nonionic detergents.
Neurotoxic esterase (NTE) is a membrane-bound protein found in highest concentration in brain and lymphocytes. The enzyme has no known physiological function, but its organophosphorylation and aging in neural tissue are thought to trigger the pathogenesis of organophosphorus-induced delayed neuropathy (OPIDN). Solubilization of NTE from microsomal membranes from hen or chick brain was studied with ten detergents encompassing ionic, zwitterionic, or nonionic types. Corrected yields of NTE solubilized over a range of [detergent]/[protein] ratios were determined by dividing the activity not sedimenting in detergent at 100,000 g for 60 min at 4 degrees by the activity in the original microsomal fraction with no detergent present. Highest corrected yields were obtained with sodium cholate (44%), Triton X-100 (48%), and nonyl-GPS (57%). Partial loss of NTE activity occurred in the presence of detergent which could be prevented by the inclusion of asolectin in the solubilization preparation. NTE could not be solubilized by omitting detergent or by substituting 2 M NaCl for detergent. Mipafox pI50 values obtained from complete titration curves carried out on NTE solubilized in Triton X-100, sodium cholate, or sodium cholate/asolectin were indistinguishable from the value for native enzyme from brain homogenate. These results indicate that NTE exhibits the properties of an integral membrane protein with lipid dependence. The enzyme can be solubilized in good yield with a variety of detergents with retention of its characteristic differential inhibition by paraoxon and mipafox, a necessary prelude to bulk purification of the enzymatically active protein.
D000818:Animals; D001921:Brain / Q000201:enzymology*; D002265:Carboxylic Ester Hydrolases / Q000302:isolation & purification / Q000378:metabolism*; D002645:Chickens; D003902:Detergents; D007425:Intracellular Membranes / Q000201:enzymology*; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology*; D012995:Solubility
D003902:Detergents; D002265:Carboxylic Ester Hydrolases; C021859:neurotoxic esterase