Characterization of high-affinity binding sites for diisopropylfluorophosphate (DFP) from chicken spinal cord membranes.
The delayed neurotoxic organophosphate [3H]diisopropylfluorophosphate ([3H]DFP) binds with high affinity to membrane-bound proteins from the chicken spinal cord. The DFP binding proteins were solubilized from membrane preparations, using a detergent (CHAPS). The protein(s) sites that labeled with a low concentration of [3H]DFP, e.g. 10(-10)-10(-9) M, were defined as the high-affinity binding sites. The density (or concentration) of the high-affinity binding sites in protein(s) was determined by the difference between total and non-specific binding. The high-affinity binding sites were saturable, and the maximal amount of binding sites was estimated at 400 fmol/mg protein. [3H]DFP binding to solubilized proteins was not completely reversible. Concentration-dependent curves suggested that the [3H]DFP binding sites differ from the active sites of acetylcholinesterase, butyrylcholinesterase, and neuropathy target esterase, as well as from muscarinic acetylcholine receptors. The amount of DFP binding sites after a neurotoxic dose of tri-o-cresyl phosphate (TOCP) decreased markedly in membrane preparations from the chicken spinal cord. These results indicate that a TOCP metabolite(s) interacts with the DFP binding sites in vivo. Gel filtration chromatography of the solubilized membranes indicated at least two major high-affinity DFP binding proteins with apparent molecular weights of 300 and 110 kDa. The DFP binding sites corresponding to the 110 kDa protein were insensitive to eserine, a potent anti-cholinesterase agent.
Konno
N
N
Department of Public Health, Fukushima Medical College, Japan.
Suzuki
N
N
Horiguchi
H
H
Fukushima
M
M
D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D002645:Chickens; D002850:Chromatography, Gel; D007531:Isoflurophate / Q000737:chemistry*; D009422:Nervous System Diseases / Q000139:chemically induced; D010830:Physostigmine / Q000494:pharmacology; D013116:Spinal Cord / Q000737:chemistry*; D014317:Tritolyl Phosphates / Q000494:pharmacology / Q000633:toxicity
D014317:Tritolyl Phosphates; D007531:Isoflurophate; D010830:Physostigmine; C025541:tri-o-cresyl phosphate