Effect of 5-Hydroxytryptamine on Sodium- and Potassium-Dependent Adenosine Triphosphatase and Its Reactivity toward Ouabain

Activity of vertebrate Na+,K+-ATPase was inhibited by 5-hydroxytryptamine. Inhibition was reversible, and activity could be restored by dilution of 5-hydroxytryptamine (5HT). The ability of indole derivatives to inhibit depended on the presence of a net charge on the third atom of the indole C-3 side chain. Indoles with a net positive charge, such as 5HT, were stronger inhibitors than those with a net negative charge. Derivatives with a net charge of zero were not inhibitors. The ability of indole derivatives to inhibit Na+,K+-ATPase activity decreased in the order 5HT > tryptamine > tryptophanamide > tryptophol >N-acetyl-5-methoxytryptamine > indole-3-acetic acid. Trp did not inhibit either ATPase or pNPPase activity. Charged indole derivatives also inhibited the pNPPase activity of Na+,K+-ATPase. 5HT and tryptophanamide were stronger inhibitors of pNPPase activity than indole-3-acetic acid. Binding of [3H]ouabain was inhibited by 5HT and tryptophanamide. Trp did not inhibit [3H]ouabain binding. The near equilibrium level of [3H]ouabain binding in the presence of ATP, Mg2+, and Na+was decreased by 5HT in the manner characteristic of a competitive inhibitor. Enzyme-bound [3H]ouabain could be displaced by 5HT. 5HT was a complex mixed inhibitor of K+with interactions at two sites, Tris was a competitive inhibitor with interactions at three sites, and ouabain was a simple noncompetitive inhibitor. The data are explained through a model indole binding site containing oppositely charged residues.