Properties of the tetrodotoxin binding component in plasma membranes isolated from Electrophorus electricus.
The biochemical properties of the electrically excitable sodium channels in the electroplaque of Electrophorus electricus were investigated using tritiated tetrodotoxin (TTX) as a specific membrane probe. Membrane fragments from the electroplaque were isolated essentially by differential centrifugation and characterized with respect to the plasma membrane markers acetylcholine receptors, acetylcholinesterase, (Na+ + K+)ATPase, and [3H]TTX binding. Equilibrium binding studies showed that [3H]TTX bound to a single population of noninteracting receptor sites with an apparent dissociation constant of 6 +/- 1 X 10(-9) M. The toxin-membrane complex dissociated with a first-order rate constant of 0.012 sec-1. Studies on the pH dependence of complex formation demonstrated the requirement for an ionizable, functional group with a pK of 5.3 and this group has been shown to be a carboxyl. Treatment of the membranes with trimethyloxonium tetrafluoroborate, a carboxyl group modifying reagent, resulted in an irreversible loss in the binding of [3H]TTX, which could be prevented by low concentrations of TTX or saxitoxin. This decrease was due to a reduction in the total number of binding sites and not to a decrease in toxin binding affinities. The relative binding affinities of various monovalent alkali metal and polyatomic cations for the TTX-receptor site showed that this site displayed cation discrimination properties which were similar to those reported previously for the electrically excitable sodium channel in intact nerve fibers. A possible role for this site in the ion selectivity of the sodium channel is proposed.
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002414:Cations, Monovalent; D004557:Electric Organ / Q000378:metabolism*; D004593:Electrophorus; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008433:Mathematics; D009928:Organ Specificity; D009994:Osmolar Concentration; D011485:Protein Binding; D011955:Receptors, Drug / Q000187:drug effects; D012965:Sodium Chloride / Q000494:pharmacology; D013045:Species Specificity; D013779:Tetrodotoxin / Q000378:metabolism*
D002414:Cations, Monovalent; D011955:Receptors, Drug; D013779:Tetrodotoxin; D012965:Sodium Chloride