Researchers identify a new role for covalent linker involved in biological ion channel

Ion channels are essentially passageways through membranes that transport signals to a cell’s environment and enable it to respond.

Jianhan Chen. Image Credit: University of Massachusetts Amherst.

In the molecular-level realm of ion channels, scientists have discussed about the function of a tiny piece of the channel known as a linker, stated computational biophysicist Jianhan Chen from the University of Massachusetts Amherst.

The linker interacts between the pore and its environment-detecting apparatus, and learning its function—that is, whether it plays an active sensing role or is inert—has not been clear. However, it might result in a novel target for drugs and treatment for conditions like autism, asthma, stroke, epilepsy, and hypertension, added Chen.

At Washington University, Chen and his collaborators have recently reported in the eLife journal that their experiments have exposed “the first direct example of how non-specific membrane interactions of a covalent linker can regulate the activation of a biological ion channel.”

Specifically, Chen and the study’s co-first authors Mahdieh Yazdani and Zhiguang Jia from the University of Massachusetts Amherst, with co-first author Guohui Zhang, Jingyi Shi, and Jianmin Cui from Washington University, examined a pore known as the large-conductance potassium (BK) channel.

The channel is significant in neuron and muscle function and is regulated by the concentration of calcium through a calcium-sensing domain. It is also regulated by electrical potential via a voltage-sensing domain. Either way, it closes or opens similar to a gate—“a really common architecture in transmembrane receptors and channels,” added Chen.

One tiny “C-linker” links the pore and the BK calcium sensor and, to date, was assumed to be mainly an inert connection. To examine it, Chen stated, “The traditional approach, if you suspect a specific position of the protein is important, is to mutate it and see what happens. You replace one amino acid with another. But with this method, you could end up perturbing many things; it’s hard to tell what you’ve done.”

The scientists instead scrambled the sequence of the C-linker amino acid several times.

If you do enough scrambles, you create so many different effects that you can average them. If the function isn’t changed and all the repetitions look basically the same, nothing will stand out. This will also give you a clean background so that next you can test some specific force or type of interaction that the linker might be involved in.”

Jianhan Chen, Computational Biophysicist, University of Massachusetts Amherst

The researchers found that scrambling the linker significantly influenced the activation of the BK channel, supporting the idea that the linker is not just an inert connection but more than that, Chen added. Unexpectedly, computational analysis had predicted that it was not the sensor or pore, but the nonspecific linker-membrane interactions that resulted in different channel properties.

Chen informed that Zhang from Washington University performed “two really elegant experiments” to test this unique mode of channel regulation. Zhang created a shorter version of the channel without the calcium sensor but without affecting its voltage-sensing function.

If our hypothesis is correct, in this construct the linker scramble would affect this truncated channel in a similar fashion as in the full-length channel. And if the linker does react without the calcium domain there, the linker is interacting with something else.”

Jianhan Chen, Computational Biophysicist, University of Massachusetts Amherst

That proved to be the case. Moreover, the researchers took one of the scrambling mutants and eliminated the “membrane-anchoring” segment that communicated with the membrane, Chen added.

“We show that this single change completely reverses the linker scrambling effect. This one particular anchoring piece is responsible for the functional differences we observed,” he further added.

In addition to improving knowledge, a significant part of the finding relates to the number of other domain-to-domain linkers present in membrane proteins, Chen explained.

He added, “Now we must really consider that the linker itself is part of the sensing apparatus, rather than just a connection. It’s a new way to think about it. Our study is a really strong argument that the linker is a lot more important than some people thought.”

We don’t discuss a direct application to disease, but this paper offers an important insight. We think it’s going to spark others to do more. It could offer a new way to design drugs because now you can also think about also targeting the linker, not only the sensing domain or the pore itself. It gives you one more possibility.”

Jianhan Chen, Computational Biophysicist, University of Massachusetts Amherst