Structure of the CLC-1 chloride channel from Homo sapiens

Abstract

CLC channels mediate passive Cl ⁻ conduction, while CLC transporters mediate active Cl ⁻ transport coupled to H ⁺ transport in the opposite direction. The distinction between CLC-0/1/2 channels and CLC transporters seems undetectable by amino acid sequence. To understand why they are different functionally we determined the structure of the human CLC-1 channel. Its ‘glutamate gate’ residue, known to mediate proton transfer in CLC transporters, adopts a location in the structure that appears to preclude it from its transport function. Furthermore, smaller side chains produce a wider pore near the intracellular surface, potentially reducing a kinetic barrier for Cl ⁻ conduction. When the corresponding residues are mutated in a transporter, it is converted to a channel. Finally, Cl ⁻ at key sites in the pore appear to interact with reduced affinity compared to transporters. Thus, subtle differences in glutamate gate conformation, internal pore diameter and Cl ⁻ affinity distinguish CLC channels and transporters.

eLife digest

Channels and transporters are two classes of proteins that transport molecules and ions – collectively referred to as “substrates” – across cell membranes. Channels form a pore in the membrane and the substrates diffuse through passively. Transporters, on the other hand, actively pump substrates across a membrane, consuming energy in the process. Thus, channels and transporters work in distinct ways.

Channels and transporters most often have unrelated structures, but there are rare examples of both existing within the same family of structurally similar proteins. CLC proteins, for example, include both chloride ion channels and transporters that pump chloride ions in one direction by harnessing the energy from hydrogen ions flowing in the other direction.

It remains unclear why some CLC proteins work as channels while others are transporters, especially since the two seem indistinguishable on the basis of the order of their amino acids – the building blocks of all proteins. The conservation of the amino acid sequences implies they are structurally very similar. How then can different members perform such energetically distinct processes?

Park and MacKinnon now show that the answer to this question serves as a reminder of how subtle nature can be. Indeed, while the structure of a human CLC channel (called CLC-1) is indeed similar to those of CLC transporters, one amino acid adopts a unique shape that explains why the protein cannot act as a transporter. This specific amino acid, a glutamate, is central to the exchange of chloride and hydrogen ions in CLC transporters. Park and MacKinnon show that its conformation in the CLC-1 channel stops this exchange, while leaving the pore open for the passive transport of chloride ions. Also, two other amino acids along the ion diffusion pathway in the CLC channel are smaller than their counterparts in CLC transporters, and so allow chloride ions to diffuse through more quickly. Lastly, Park and MacKinnon also note that channels do not require a wide pore: instead ions can still flow rapidly through a narrow pore if the chemical environment inside permits it.

CLC proteins perform a number of important roles in humans, and mutations in CLC-encoding genes underlie numerous heritable diseases. It remains too early to know how this mechanistic study may or may not impact treatments, yet the findings will likely interest scientists working on ion conduction mechanisms and the evolution of molecular function.