Catalytic Activity Of Acetylcholinesterase Research Articles

Synthesis and reaction of some S-alkyl diphenylthiophosphinates with acetylcholinesterase

1. A number of S-alkyl diphenylthiophosphinates was synthesized and their inhibiting activity toward acetytcholinesterase was studied. 2. The obtained compounds are capable of both reversible and irreversible inhibition of the catalytic activity of acetylcholinesterase.

Effect of α-Bungarotoxin on Acetylcholinesterase bound to Mouse Diaphragm Endplates

IT has been shown that α-bungarotoxin (α-Bgt) irreversibly blocks cholinoreceptors1–10, and that D-(+)tubocurarine (TC) protects these receptors from the toxin4–7. On the other hand, it has been emphasized that α-Bgt has no effect on the catalytic activity of acetylcholinesterase (AChE)4,6,9, and it was concluded that the cholinoreceptor and AChE must be two different macromolecules8,9. Because typical cholinolytics, including TC and gallamine, characteristically influence the kinetics of a membrane-bound AChE11,12, it seemed justified to reinvestigate whether α-Bgt really is an exception in this respect.

In Vitro excitation of purified membrane fragments by cholinergic agonists

Excitation of membrane fragments by cholinergic agonists is measuredin vitro by a filtration technique. Membrane fragments which contain high levels of the enzyme acetylcholinesterase and presumably originate from the innervated excitable faces of electroplax are first purified from homogenates of electric organ ofElectrophorus electricus by centrifugation in a sucrose gradient. Then the fragments, which make closed vesicles or microsacs, are equilibrated overnight with a medium containing(22)Na(+). After equilibration of the inside of the microsacs with the outside medium, the suspension is diluted into a nonradioactive medium. The(22)Na(+) content of the microsacs as a function of time is then followed by rapid filtration on Millipore filters. In the presence of cholinergic agonists, the time course of(22)Na(+) release changes: the rate of(22)Na(+) release increases. This increase is blocked byd-tubocurarine and is absent with microsacs derived from the non-innervated inexcitable membrane of the electroplax. The response to cholinergic agonists is thus followed on a completely cell-free system, in a well-defined environment. The dose-response curves to cholinergic agents obtainedin vitro agree, quantitatively, with the dose-response curves recordedin vivo by electrophysiological methods. In particular, the dose-response curve to agonists is sigmoid, the antagonism betweend-tubocurarine and carbamylcholine competitive, and the antagonism between tetracaine and carbamylcholine noncompetitive. The effects of two different affinity labeling reagents on the response to agonists and on the catalytic activity of acetylcholinesterase are followed in parallel on the same microsac preparation. The effects of dithiothreitol and of gramicidin A on the microsacs are studied and are found to be similar to those observedin vivo with the isolated electroplax.