2. Academic Validation
  3. Mechanism of Gating and Isoform-Specific Inhibition in Renal CLC Chloride Channels

Mechanism of Gating and Isoform-Specific Inhibition in Renal CLC Chloride Channels

  • bioRxiv. 2026 Feb 18:2026.02.17.706469. doi: 10.64898/2026.02.17.706469.
Chih-Ta Chien 1 2 3,  Briana L Sobecks 1 4 5 6 7 8,  Alexander S Powers 1 4 6 7 8 9,  Jürgen Kreiter 4 10,  Anindita Das 4,  Chloe N Barry 11,  Muyuan Chen 12,  Andrew Hinman 11,  Camille F Petrakian 6 7 8 9,  Natasa Trifkovic 4,  Brianna Williams 11,  Chase A P Wood 4,  Mengyuan Xu 4,  Ron O Dror 5 6 6 8,  Wah Chiu 2 12 13,  Merritt Maduke 4
Affiliations

Affiliations

  • 1 co-first authors.
  • 2 Department of Bioengineering, Stanford University, Stanford, CA 94305.
  • 3 current affiliation: National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), National Institutes of Health, Bethesda, MD, 20892.
  • 4 Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305.
  • 5 Department of Chemical Engineering, Stanford University, Stanford, CA 94305.
  • 6 Department of Computer Science, Stanford University, Stanford, CA 94305.
  • 7 Department of Structural Biology, Stanford University, Stanford, CA 94305.
  • 8 Institute for Computational and Mathematical Engineering, Stanford University, Stanford, CA 94305.
  • 9 Department of Chemistry, Stanford University, Stanford CA.
  • 10 current affiliation: Institute for Medical Physics and Biophysics, University of Veterinary Medicine Vienna, 1210 Vienna, Austria.
  • 11 Innovative Medicines Accelerator, Stanford University, Stanford, CA 94305.
  • 12 Division of CryoEM and Bioimaging, SSRL, SLAC National Accelerator Laboratory, Stanford University, Menlo Park 94025.
  • 13 Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305.
PMID: 41756880 DOI: 10.64898/2026.02.17.706469
Abstract

Hyponatremia is a prevalent disorder marked by excess water retention and substantial morbidity, motivating interest in the CLC-Ka Chloride Channel as a therapeutic target. Selectively inhibiting CLC-Ka without affecting the closely related CLC-Kb is essential for preventing serious side effects. However, developing isoform-selective inhibitors has been challenging because most small molecules do not distinguish between CLC-Ka and CLC-Kb, and the basis for selectivity in the few known exceptions remains unclear. The small molecule BIM1 preferentially inhibits CLC-Ka over CLC-Kb, providing an opportunity to dissect isoform-specific pharmacology. To investigate this mechanism, we determined cryo-EM structures of BIM1 and BIM15, a related nonselective analog, bound to a CLC-K variant engineered to match the human CLC-Ka binding pocket. Structural and computational analyses reveal that inhibition and isoform selectivity are anchored by interactions with a conserved lysine, with surrounding binding-site residues subtly tuning the local electrostatic environment to promote or disfavor these contacts. These analyses further identify a dynamic extracellular loop that intermittently occludes the access pathway, indicating its role as a gate for ions and inhibitors. BIM15 engages this gating loop more extensively than BIM1, suggesting that differential loop engagement contributes to inhibitor selectivity. To probe how gating reshapes this region, we solved the structure in the presence of CA2+, which favors channel opening, and found the gating loop ordered and withdrawn from the pathway. Together, these findings elucidate how CLC-K channels gate and how subtle binding-site differences and loop dynamics shape isoform-specific drug binding, providing a foundation for designing next-generation CLC-Ka inhibitors.

Keywords

CLC-Ka chloride channel; Major: Biological Sciences; Minor: Biophysics and Computational Biology; channel gating; cryo-EM; isoform selectivity; molecular dynamics simulations.

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