Structural basis for the ion selectivity of potassium-chloride cotransporter KCC4 revealed by cryo-EM titrationOA
Potassium-chloride cotransporters KCCs mediate the coupled,electroneutral cotransport of K^(+)and Cl^(-)across the membrane and are involved in important physiological processes such as cell volume regulation andγ-aminobutyric acid(GABA)and glycine-mediated inhibitory neurotransmission.Although structures of KCCs have been reported,the identification of ions bound in KCCs awaits experimental studies.Here using the cryo-electron microscopy(cryo-EM)titration methods,we present six structures of human KCC4 in different ion conditions at 2.38–2.58Åresolutions.These structures,along with molecular dynamic simulations,allow us to assign one K^(+)and two Cl^(-)ions in the substratebinding pocket.The K^(+)at S1 and Cl^(-)at the S2 site are tightly coupled in the binding and dissociation,suggesting that the Cl^(-)at S2 but not at S3 is the cotransported one.The S1 site provides coordination that largely matches the K^(+)dehydration radius and therefore displays higher selectivity to K^(+)over Na^(+).This study establishes the structural basis for the K^(+)selectivity of KCCs by the cryo-EM titration.
Yuan Xie;Binming Han;Xin Tao;Fangjun Song;Cheng Zhao;Eric Delpire;Jingyuan Li;Shan Wu;Jiangtao Guo
Department of Neurosurgery,Xijing Hospital,Fourth Military Medical University,Xi''an 710032,ChinaSchool of Physics,Zhejiang University,Hangzhou 310058,ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering,Hubei Collaborative Innovation Center for Green Transformation of Bio-Resources,Hubei Key Laboratory of Industrial Biotechnology,School of Life Sciences,Hubei University,Wuhan 430062,ChinaDepartment of Biophysics and Department of Neurology of the Fourth Affiliated Hospital,Zhejiang University School of Medicine,Hangzhou 310058,ChinaDepartment of Biophysics and Department of Neurology of the Fourth Affiliated Hospital,Zhejiang University School of Medicine,Hangzhou 310058,ChinaDepartment of Anesthesiology,Vanderbilt University School of Medicine,Nashville,TN 37232,USASchool of Physics,Zhejiang University,Hangzhou 310058,ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering,Hubei Collaborative Innovation Center for Green Transformation of Bio-Resources,Hubei Key Laboratory of Industrial Biotechnology,School of Life Sciences,Hubei University,Wuhan 430062,ChinaDepartment of Biophysics and Department of Neurology of the Fourth Affiliated Hospital,Zhejiang University School of Medicine,Hangzhou 310058,China Nanhu Brain-Computer Interface Institute,Hangzhou 311100,China NHC and CAMS Key Laboratory of Medical Neurobiology,MOE Frontier Science Center for Brain Science and Brain-machine Integration,School of Brain Science and Brain Medicine,Zhejiang University,Hangzhou 310058,China State Key Laboratory of Plant Environmental Resilience,College of Life Sciences,Zhejiang University,Hangzhou 310058,China Cancer Center,Zhejiang University,Hangzhou 310058,China
生物科学
KCC4Cryo-EM titrationIons selectivityMolecular dynamic simulationIons cotransport
《Biophysics Reports》 2026 (3)
P.193-206,14
supported in part by the National Natural Science Foundation of China(32200803 to Y.X.,32371204 to J.G.,32471319 to S.W.,and 11722434 and 11874319 to J.L.)the Ministry of Science and Technology(2020YFA0908501 to J.G.)the Fundamental Research Funds for the Central Universities(2021FZZX001-28 and 2025ZFJH01-01 to J.G.)supported MOE Frontier Science Center for Brain Science&Brain-Machine Integration,Zhejiang Universitythe support of K.C.Wong Education Foundationgranted by the A-level Doctoral Research Foundation from Xijing Hospital,Fourth Military Medical University(XJARC05)by Special Project of A-level Doctoral Talent Support Program from Xijing Hospital,Fourth Military Medical University(AL202406)supported by the Distinguished Young Scholars of Hubei Province(2022CFA078)the Knowledge Innovation Program of the Wuhan-Shugung Project(2023020201020418)the National Key Research and Development Program of China(2020YFA0908400)。
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