Further investigations on a Mg++ + Na+-activated adenosintriphosphatase, possibly related to the active, linked transport of Na+ and K+ across the nerve membrane

Abstract

The sodium activated ATPase prepared from crab nerve which was described in a previous paper1 has been subjected to a more detailed study of the relation between enzyme, substrate, and a number of monovalent and divalent cations. It has also been investigated whether the enzyme catalyses an ATP-Pi or an ATP-ADP exchange reaction, and whether these reactions are cation-dependent. Finally, the effect of g-strophanthin on the ATPase activity and the exchange reaction has been investigated. The simplest scheme by which the experimental results can be explained is as follows: (1)E + ATP + 2 Mg + nNa + mK ⇌ E nNa mKMg2ATP(2) E nNa mKMg2ATP ⇌ E nNa mK ∼ 2 Mg + ADP(3) E nNa mK ∼ P → E + Pi + nNa + mK where n and m are unknown numbers, and where E nNa mKMg2ATP symbolises an enzyme-substrate-ion complex in which Mg++ is part of the enzyme-substrate complex and in which Na+ and K+ are attached to two different sites on the enzyme; these sites show high affinities for the respective ions. Phosphorylation of the enzyme, steps (1) + (2), will take place also when Mg++ is the only cation present in the system, but under these circumstances the release of inorganic phosphate, step (3), is very slow. The rate of step (3) is increased by the addition of Na++ to the system, but not by the addition of K+. In the presence of Mg++ + Na+, however, addition of K+ leads to a considerable increase of the rate of step (3). The binding of Na+ + K+ to the enzyme takes place during the attachment of ATP to the enzyme, i.e. during step (1). G-strophanthin has no effect on the phosphorylation of the enzyme, steps (1) + (2). But g-strophanthin inhibits the activating effect of Na+ and of Na+ + K+ on step (3), presumably by interfering with the binding of the cations to the enzyme during step (1). The observations lend further support to the suggestion made previously that this enzyme is involved in the active, linked transport of Na+ and K+ across the nerve membrane. © 1960.