NaK ATPase Maintains the Intracellular Na and K Concentrations in Animal Cells

A second important P-class ion pump present in the plasma membrane of all animal cells is the Na+/K+ ATPase. This ion pump is a tetramer of subunit composition a2p2. (Classic Experiment 7.1 describes the discovery of this enzyme.) The small, glycosylated p polypeptide helps newly synthesized a subunits to fold properly in the endoplasmic reticulum but apparently is not involved directly in ion pumping. The amino acid sequence and predicted secondary structure of the catalytic a subunit are very similar to those of the muscle SR Ca2+ ATPase (see Figure 7-8). In particular, the Na+/K+ ATPase has a stalk on the cytosolic face that links

▲ FIGURE 7-9 Operational model of the Na+/K+ ATPase in the plasma membrane. Only one of the two catalytic a subunits of this P-class pump is depicted. It is not known whether just one or both subunits in a single ATPase molecule transport ions. Ion pumping by the Na+/K+ ATPase involves phosphorylation, dephosphorylation, and conformational changes similar to those in the muscle

Ca2+ ATPase (see Figure 7-7). In this case, hydrolysis of the E2-P Intermediate powers the E2 n E1 conformational change and concomitant transport of two ions (K+) inward. Na+ ions are indicated by red circles; K+ ions, by purple squares; high-energy acyl phosphate bond, by ~P; low-energy phosphoester bond, by -P [See K. Sweadner and C. Donnet, 2001, Biochem. J. 356:6875, for details of the structure of the a subunit.]

domains containing the ATP-binding site and the phospho-rylated aspartate to the membrane-embedded domain. The overall transport process moves three Na+ ions out of and two K+ ions into the cell per ATP molecule hydrolyzed.

The mechanism of action of the Na+/K+ ATPase, outlined in Figure 7-9, is similar to that of the muscle calcium pump, except that ions are pumped in both directions across the membrane. In its E1 conformation, the Na+/K+ ATPase has three high-affinity Na+-binding sites and two low-affinity K+-binding sites accessible to the cytosolic surface of the protein. The Km for binding of Na+ to these cytosolic sites is 0.6 mM, a value considerably lower than the intracellular Na+ concentration of =12 mM: as a result, Na+ ions normally will fully occupy these sites. Conversely, the affinity of the cytosolic K+-binding sites is low enough that K+ ions, transported inward through the protein, dissociate from E1 into the cytosol despite the high intracel-lular K+ concentration. During the E1 n E2 transition, the three bound Na+ ions become accessible to the exoplasmic face, and simultaneously the affinity of the three Na+-binding sites becomes reduced. The three Na+ ions, transported outward through the protein and now bound to the low-affinity Na+ sites exposed to the exoplasmic face, dissociate one at a time into the extracellular medium despite the high extracellular Na+ concentration. Transition to the E2 conformation also generates two high-affinity K+ sites accessible to the exoplasmic face. Because the Km for K+ binding to these sites (0.2 mM) is lower than the extracellular K+ concentration (4 mM), these sites will fill with K+ ions. Similarly, during the E2 n E1 transition, the two bound K+ ions are transported inward and then released into the cytosol.

Certain drugs (e.g., ouabain and digoxin) bind to the exoplasmic domain of the plasma-membrane Na+/K+ ATPase and specifically inhibit its ATPase activity. The resulting disruption in the Na+/K+ balance of cells is strong evidence for the critical role of this ion pump in maintaining the normal K+ and Na+ ion concentration gradients.

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