Effect of Organophosphorus Compounds on the Conformation of Acetylcholinesterase and Acetylcholine Receptor. Reconstitution of Globular Dimer of Acetylcholinesterase and Interaction of Acetylcholinesterase and Receptor with Diisopropylfluorophosphate
Annual rept. 15 Feb 1986-14 Feb 1987
CALIFORNIA UNIV SAN FRANCISCO
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The detergent-soluble globular dimer of acetylcholinesterase from Torpedo californica was reconstituted through dialysis into egg phosphatidylcholine vesicles. The size of the reconstituted particles depended on the ionic strength of the buffer as well as the molar lipidprotein ration R. The solution of the protein-lipid complex was turbid at R 5,000 and I 0.13, and the particles became heterogeneous at R 2,000. The enzyme was unstable at R 1,000 and I 0.05. Based on circular dichroism studies, the conformation of the enzyme reconstituted at R 4,000 and I 0.07 remained unaltered. The enzymatic activity and the Michaelis-Menten constant were also unchanged. The reconstituted enzyme seemed to be more stable against thermal denaturation than in detergent solution. Acetylcholinesterase is irreversibly inhibited by diisopropyl fluorophosphate DFP. The three isozymes, buffer- soluble globular dimer, asymmetric dodecamer and its derived globular tetramer, had essentially the same bimolecular rate constant. The acetylcholine receptor from Torpedo californica had virtually the same conformation in the absence and presence of DFP. Its binding affinity for alpha-bungarotoxin was also unaffected by the addition of DFP. Initial study of sodium flux through the receptor suggested that DFP might affect, in a slow, time-dependent manner, the accumulation of sodium ions. Keywords sharks.