title: Characterization of [3H]di-isopropyl phosphorofluoridate-binding proteins in hen brain. Rates of phosphorylation and sensitivity to neurotoxic and non-neurotoxic organophosphorus compounds.
abstract: The experiments described in this paper were designed to isolate [3H]di-isopropyl phosphorofluoridate-binding proteins by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis for the purpose of characterizing and identifying potential initiation sites for organophosphorus-compound-induced delayed neurotoxicity. The major Paraoxon-insensitive Mipafox-sensitive binding protein (Mr 160 000) was found to be identical with one previously identified as neurotoxic esterase, an enzyme that has been proposed to be the target site for organophosphorus-compound-induced delayed neurotoxicity. However, two other binding proteins with suitable binding characteristics were also found in smaller amounts, one of which has not been detected previously. Di-isopropyl phosphorofluoridate was found to phosphorylate all three of these proteins at rates similar to the rate at which neurotoxic esterase is inhibited by di-isopropyl phosphorofluoridate. Varying the concentration of di-isopropyl phosphorofluoridate or the time of incubation produced similar increases in binding to each of the labelled proteins. This suggests that the reaction rates of di-isopropyl phosphorofluoridate with proteins may be described by first-order kinetics, and the concentration of the Michael is complex formed during binding is minimal for all the phosphorylated proteins. The recovery of the binding activity in the 160 000-Mr band was found to be similar to the recovery of neurotoxic esterase activity, lending further support to the contention that this band is identical with neurotoxic esterase.
mesh_terms: D000818:Animals; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D007531:Isoflurophate / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D008565:Membrane Proteins / Q000378:metabolism; D010261:Paraoxon / Q000494:pharmacology; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins; D011506:Proteins; D007531:Isoflurophate; C005238:mipafox; D010664:Phenylmethylsulfonyl Fluoride; D010261:Paraoxon