Transporting ions across cellular membranes depends on the activity of little pumps that are of fundamental importance to life. They utilize the energy derived from ATP hydrolysis to establish and maintain electrochemical ion gradients across membranes that are the basis of vital processes such as nutrient uptake, pH homeostasis or electrical excitability. A significant number of them belong to the family of P-type ATPases, named for the formation of a phosphorylated intermediate in the reaction cycle. Although closely related, different P-type ATPases transport different ions. An unsolved question so far is how these pumps discriminate between different ions, and a recent Nature paper describing the atomic structure of the Na+,K+-ATPase, determined by a team of Danish scientists led by Bente Vilsen and Poul Nissen, sheds new light on ATPase ion discrimination.
The Na+,K+-ATPase is a transmembrane enzyme that transports three sodium ions for every two potassium ions pumped in the opposite direction. In contrast to more simple P-type ATPases, such as SERCA(a Ca2+ pump composed of a single subunit), the Na+,K+-pump is made of three subunits. Itsα-subunit shares significant homology with SERCA, while the β- andγ-subunits are unique to the Na+,K+-ATPase. According to the classical model of the ion pump reaction cycle, P-type ATPases adopt two conformations, known as E1 and E2. During the reaction cycle the transported sodium and potassium ions are temporarily trapped in the protein's E1 and E2 state, respectively, ensuring that the entry gate closes before the exit gate opens, a mechanism that prevents leakiness. But does this classical model really explain how the Na+,K+-ATPase functions?
To get an initial snapshot of the Na+,K+-ATPase, the Danish scientists purified the enzyme from pig kidneys and crystallized it in the presence of rubidium ions, which behave like potassium ions but are large enough to be visualized in the crystal structure. The team expected that their snapshot would provide a view of the Na+,K+-ATPase with two rubidium ions trapped in the same location as the potassium ions during the pumping process.
Analysis of the Na+,K+-ATPase structure shows that the β- and γ-subunits interact with three of the α-subunit's helices in the transmembrane region. Interestingly, the carboxy-terminal end of the α-subunit contains positively charged arginine residues, which may act as membrane voltage sensors. As expected, the two rubidium ions were trapped in the transmembrane part of the α-subunit, confirming the classical model of the reaction cycle. Surprisingly, the α-subunit had an unexpectedly high structural similarity to SERCA, even in the binding pocket, posing the question of how such similar proteins distinguish between the different ions that they pump. The team concluded that subtle differences in side chain and water molecule positions must be sufficient to determine ion selectivity.
By providing the first structural model of the Na+,K+-ATPase, Vilsen and Nissen's team are following in the footsteps of Jens Christian Skou, who ultimately earned the Nobel Prize after discovering this fundamental pump. However, to completely understand how Na+,K+-ATPases work, we will need further structures showing the enzyme in different states to give us a detailed picture of this remarkable protein's function.
