Bottom-up design of Ca2+ channels from defined selectivity filter geometry

• 成果类型: Article
• 署名作者: Liu, Yulai; Weidle, Connor; Mihaljevic, Ljubica; Watson, Joseph L.; Li, Zhe; Yu, Le Tracy; Majumder, Sagardip; Borst, Andrew J.; Carr, Kenneth D.; Kibler, Ryan D.; Gamal El-Din, Tamer M.; Catterall, William A.; Baker, David
• 署名单位: University of Washington; University of Washington Seattle; University of Washington; University of Washington Seattle; University of Washington; University of Washington Seattle; University of Washington; Howard Hughes Medical Institute; University of Washington Seattle; University of Washington; University of Washington Seattle
• 刊物名称: NATURE
• ISSN/ISSBN: 0028-0836; 1476-4687
• DOI: 10.1038/s41586-025-09646-z
• 发表日期: 2025-12-11
• 关键词: METAL-LIGAND INTERACTIONS; ion permeation; calcium-channels; structural basis; voltage; pore; inhibition; mechanism; relevant; locus

摘要

Native ion channels play key roles in biological systems, and engineered versions are widely used as chemogenetic tools and in sensing devices1,2. Protein design has been harnessed to generate pore-containing transmembrane proteins, but the design of selectivity filters with precise arrangements of amino acid side chains specific for a target ion, a crucial feature of native ion channels3, has been constrained by the lack of methods for placing the metal-coordinating residues with atomic-level precision. Here we describe a bottom-up RFdiffusion-based approach to construct Ca2+ channels from defined selectivity filter residue geometries, and use this approach to design symmetric oligomeric channels with Ca2+ selectivity filters having different coordination numbers and different geometries at the entrance of a wider pore buttressed by multiple transmembrane helices. The designed channel proteins assemble into homogeneous pore-containing particles and, for both tetrameric and hexameric ion-coordinating configurations, patch-clamp experiments show that the designed channels have higher conductances for Ca2+ than for Na+ and other divalent ions (Sr2+ and Mg2+) that are eliminated after mutation of selectivity filter residues. Cryogenic electron microscopy indicates that the design method has high accuracy: the structure of the hexameric Ca2+ channel is nearly identical to that of the design model. Our bottom-up design approach now enables the testing of hypotheses relating filter geometry to ion selectivity by direct construction, and provides a roadmap for creating selective ion channels for a wide range of applications.