COMPARED EFFECTS OF DITHIOTHREITOL ON THE INTERACTION OF AN AFFINITY-LABELING REAGENT WITH ACETYLCHOLINESTERASE AND THE EXCITABLE MEMBRANE OF THE ELECTROPLAX*
Abstract
p-(trimethyl ammonium) benzene diazonium difluoroborate (TDF), an affinity-labeling reagent of the acetylcholine receptor site(s), which in the normal cell acts as an irreversible inhibitor becomes a reversible activator after in vivo exposure of the electroplax to dithiothreitol (DTT), a disulfide bond reducing agent. After in vitro exposure of acetylcholinesterase to DTT, TDF becomes a reversible competitive inhibitor of the enzyme, using indophenyl acetate as the substrate. Both acetylcholinesterase and the macromolecular receptor of acetylcholine thus contain disulfide bonds. Additional experiments with DTT suggest that there might exist several different classes of receptor sites for cholinergic agents in the excitable membrane of the electroplax.
Footnotes
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↵ * The Hill coefficient (nH) of the response curve to Deca immediately following DTT treatment is close to 2 as in the control, whereas the nH of the response to Carb is reduced to 1. However, 30 min after DTT treatment the nH of the residual response to Deca decreases as well, to a value close to 1. The apparent depolarizing effect of TDF is due to TDF and not to a product of the reaction of TDF with DTT: if the native electroplax is exposed to a mixture of TDF and DTT, no depolarization occurs.
