9O95

Cryo-EM structure of CLC-ec1 at pH 7.5

Summary for 9O95

• Entry DOI: 10.2210/pdb9o95/pdb
• EMDB information: 70242
• Descriptor: H(+)/Cl(-) exchange transporter ClcA, CHLORIDE ION (2 entities in total)
• Functional Keywords: transmembrane protein cryo-em antiporter, transport protein
• Biological source: Escherichia coli
• Total number of polymer chains: 2
• Total formula weight: 104372.53

Authors

Chien, C.-T.,Chiu, W.,Maduke, M. (deposition date: 2025-04-17, release date: 2026-01-07, Last modification date: 2026-02-18)

Primary citation

Aydin, D.,Chien, C.T.,Kreiter, J.,Nava, A.R.,Portasikova, J.M.,Fojtik, L.,Sobecks, B.L.,Mosquera, C.,Man, P.,Dror, R.O.,Chiu, W.,Maduke, M.
Molecular mechanism of exchange coupling in CLC chloride/proton antiporters.
Nat Commun, 17:1342-1342, 2026

PubMed Abstract: The ubiquitous CLC membrane transporters are unique in their ability to exchange anions for cations. Despite extensive study, there is no mechanistic model that fully explains their 2:1 Cl/H stoichiometric exchange mechanism. Here, we provide such a model. Using differential hydrogen-deuterium exchange mass spectrometry, cryo-EM structure determination, and molecular dynamics simulations, we uncovered conformational dynamics in CLC-ec1, a bacterial CLC homolog that has served as a paradigm for this family of transporters. Simulations based on a cryo-EM structure at pH 3 revealed critical steps in the transport mechanism, including release of Cl ions to the extracellular side, opening of the inner gate, and water wires that facilitate H transport. Surprisingly, these water wires occurred independently of Cl binding, prompting us to reassess the relationship between Cl binding and Cl/H coupling. Using isothermal titration calorimetry and quantitative flux assays on mutants with reduced Cl binding affinity, we conclude that, while Cl binding is necessary for coupling, even weak binding can support Cl/H coupling. By integrating our findings with existing literature, we establish a complete and efficient CLC 2:1 Cl/H exchange mechanism.

PubMed: 41507156
DOI: 10.1038/s41467-025-68098-1

Experimental method

ELECTRON MICROSCOPY (3.2 Å)