New view of a familiar ion pump answers key questions about how it works

For some, the sodium-potassium pump may be an old acquaintance. It is a remarkably well-studied protein, discovered in the 1950s by Danish researcher Jens Christian Skou – a discovery that earned him the Nobel Prize in Chemistry in 1997. Yet despite decades of research, important aspects of how the pump works at the molecular level have remained unresolved, including key questions related to its vital role in the cell.

But what exactly makes this pump so important?

Mads Eskesen Christensen is a postdoc at the Department of Molecular Biology and Genetics and, together with fellow postdoc Michael Habeck, has led the project behind the scientific paper describing the new findings.

“The sodium-potassium pump is a vital protein found in all of our cells. Put simply, the protein transports sodium ions out of the cell and potassium ions into the cell across the cell membrane, thereby creating essential concentration gradients. One of the things the pump is best known for is establishing the electrochemical potentials in our neurons that allow our nervous system to function and our nerve cells to transmit signals. But these ion gradients also drive a wide range of other transport processes in cells,” explains Mads Eskesen Christensen.

In other words, the protein is essential for the body to function. Or, as Mads Eskesen Christensen puts it very simply:

“It has to work.”

A familiar pump seen in a new way

Much has happened since Jens Christian Skou discovered the pump in the 1950s, but researchers’ interest in this important protein is far from over.

When researchers have previously studied the structure of the sodium-potassium pump, they have generally “locked” the protein in specific, stable states. This has made it possible, among other things, to investigate what the pump looks like when it binds sodium or potassium by saturating it with one ion or the other.

However, this approach has its limitations. The protein is dynamic and moves through a series of different conformations as it works, and not all of these states can be captured in this way. As a result, some stages of the pump’s transport cycle have until now been impossible to study.

Using cryo-EM, the researchers have now been able to study the pump under active transport conditions. Through extensive data analysis, they were able to sort observations of the pump into different functional states and thereby uncover previously uncharacterised stages.

“Using cryo-EM, we determined protein structures of the Na,K-pump during active transport. Combined with a biochemical approach, this has allowed us to gain new insight into several of the different conformations the pump adopts while cyclically transporting ions across the membrane. One of the newly identified states in particular has long been highly sought after, because it represents the critical transition in the transport mechanism where the pump shifts from an inward-facing to an outward-facing state and can therefore release Na+ ions outside the cell after they have been bound inside the cell.

“This state contains important information about how the protein works, and we can relate it to the coordination chemistry that regulates the sodium-binding sites and shifts from stabilising to destabilising sodium binding,” explains Mads Eskesen Christensen.

The researchers also compared the general variant of the sodium-potassium pump, which is found in cells throughout the body, with a variant found primarily in nerve cells.

They look alike, but behave differently

The researchers had expected to find small but distinct structural differences between the two variants. However, when they examined them more closely, the individual conformations turned out to be surprisingly similar.

Instead, the difference lay in which states the two variants preferentially occupy.

“By comparing the general and neuronal variants, we found that their structures in the different states were highly similar, but that their distribution across these states under the same starting conditions was very different. We have shown that this difference in state distribution can explain how sodium binding occurs at different sodium concentrations in the two variants.

“The protein is therefore not simply a switch that can be turned on or off. Its activity is also influenced by the probability of the pump being in specific active states. By expressing several variants of the protein, our nerve cells can therefore be fine-tuned to meet specific needs, and this is essential for a well-functioning nervous system,” explains Mads Eskesen Christensen.

The new insights into the pump’s different states have also allowed the researchers to better understand what happens when a disease-related mutation alters the neuronal variant of the pump.

They found that the mutation affects the pump’s ability to bind a particular lipid in the cell membrane that helps stimulate its activity.

“When the mutation occurs, the protein can no longer bind a specific type of lipid in the cell membrane that stimulates the pump’s activity. This means that the sodium-potassium pump becomes less active, which may be part of the explanation for the disease,” explains Mads Eskesen Christensen.

At the same time, the finding may point to a mechanism that extends beyond the specific mutation studied by the researchers.

“What is interesting is that this mechanism may be more general. We see mutations in the same region not only in the sodium-potassium pump, but also in other ion pumps. It may therefore be a mechanism that is involved in several diseases.”

Basic research with a long-term perspective

“This is basic research,” Mads Eskesen Christensen emphasises.

There is therefore still a long way from these new findings to a specific treatment. However, a better understanding of how the sodium-potassium pump and its different variants function could eventually be important for developing treatments that target one specific variant without affecting the others.

For example, the goal could be to target a disease-affected variant of the pump more precisely, while allowing the healthy sodium-potassium pumps elsewhere in the body to continue carrying out their vital work.