TY - JOUR T1 - Evidence for Two Forms of Fluoride-Treated Sodium- and Potassium-Dependent Adenosine Triphosphatase JF - Molecular Pharmacology JO - Mol Pharmacol SP - 149 LP - 158 VL - 8 IS - 2 AU - STANLEY C. PENZOTTI, JR. AU - ELWOOD TITUS Y1 - 1972/03/01 UR - http://molpharm.aspetjournals.org/content/8/2/149.abstract N2 - The (Na+ + K+)-dependent ATPase (EC 3.6.1.3) from rat kidney microsomal fraction was inactivated by 5 mM Mg++ and 10 mM F- after incubation for about 1 hr at pH 8.5, but lower pH values or the presence of 30 mM KCl enhanced the enzyme activity. The inhibited enzyme was not appreciably reactivated by repeated washing of the particles with 0.1 M Tris buffer, pH 7.4, at 0°, or by incubation with buffer at 37°. Regeneration rates measured in the presence of Na+ rose with increasing concentrations of this cation and were greatly enhanced by the inclusion of 15 mM Tris-ATP in the incubation medium. Preparations which had been inactivated by F- in the presence of 30 mM K+ or had been temporarily exposed to K+ after prior inactivation recovered activity at about twice the rate of inhibited particles that had never been exposed to K+. Thus, after 10 min at 37° in 30 mM Na+ and 15 mM Tris-ATP, particles originally inhibited in the presence of 30 mM K+ regained 54% of the activity of controls, and preparations inhibited by NaF alone regained 22%. When 30 mM K+ was added to the solution of 30 mM Na+ and 15 mM ATP in which regeneration was attempted, no reactivation of the enzyme occurred. The data suggest that K+ converts the inactivated enzyme to a K+ complex that resists reactivation, and that this reaction is associated with a change in conformation of the enzyme. After this change in conformation, the enzyme-potassium complex would appear to have a dissociation constant of the order of 2 x 10-4 M. The affinity for potassium is thus sufficiently low so that washing of the particulate enzyme would be expected to remove this cation. The enzyme is then left in a form that regenerates rapidly with Na+ and ATP. ER -