Inhibitory characteristics of 3,5-dibromo-1-acetoxy-4-oxo-2,5-cyclohexadien-1-acetonitrile, a semisynthetic derivative of aeroplysinin-1 from sponges (Aplysinidae), on Na⁺ - K⁺-ATPase

3,5-Dibromo-1-acetoxy-4-oxo-2,5-cyclohexadien-1-acetonitrile (dienone A) inhibited Na⁺ - K⁺-ATPase with a half-maximal inhibition concentration (i₅₀) equal to 2.9 × 10^-6 M. Inhibition was time- and pH-dependent and complete after 20-30 min preincubation within a range of pH from 7.0 to 9.0. Kinetic evaluation of the cationic substrate activation of Na⁺ - K⁺-ATPase indicated mixed type inhibition with regard to Na⁺ and K⁺ and competitive inhibition with regard to ATP activation of the enzyme. The presence of Mg²⁺ caused an increased inhibition. Also, K⁺-p-nitrophenyl phosphatase activity was altered by dienone A and mixed type inhibition with regard to p-nitrophenyl phosphate and K⁺ was demonstrated. Inhibition was partially restored by repeated washing. Preincubation with sulfhydryl reagents protected the enzyme from inhibition. A significant linear correlation between reactive enzyme sulfhydryl contents [SH] and Na⁺ - K⁺-ATPase activity in the presence of varying concentrations of dienone A was observed. One of the factors causing cytotoxic activity of this compound might be its interaction with some thiol groups of the membrane-bound Na⁺ - K⁺-ATPase.