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- EMDB-10704: Structure of Human Potassium Chloride Transporter KCC3b (S45D/T94... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-10704 | |||||||||
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Title | Structure of Human Potassium Chloride Transporter KCC3b (S45D/T940D/T997D) in KCl | |||||||||
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Function / homology | ![]() Defective SLC12A6 causes agenesis of the corpus callosum, with peripheral neuropathy (ACCPN) / ammonium import across plasma membrane / potassium ion transmembrane transporter activity / potassium:chloride symporter activity / Cation-coupled Chloride cotransporters / chloride ion homeostasis / cellular hypotonic response / ammonium channel activity / cellular hypotonic salinity response / potassium ion homeostasis ...Defective SLC12A6 causes agenesis of the corpus callosum, with peripheral neuropathy (ACCPN) / ammonium import across plasma membrane / potassium ion transmembrane transporter activity / potassium:chloride symporter activity / Cation-coupled Chloride cotransporters / chloride ion homeostasis / cellular hypotonic response / ammonium channel activity / cellular hypotonic salinity response / potassium ion homeostasis / cell volume homeostasis / potassium ion import across plasma membrane / monoatomic ion transport / chloride transmembrane transport / potassium ion transmembrane transport / cellular response to glucose stimulus / chemical synaptic transmission / basolateral plasma membrane / angiogenesis / axon / synapse / protein kinase binding / membrane / metal ion binding / plasma membrane Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.08 Å | |||||||||
![]() | Chi G / Man H / Ebenhoch R / Reggiano G / Pike ACW / Wang D / McKinley G / Mukhopadhyay SMM / MacLean EM / Chalk R ...Chi G / Man H / Ebenhoch R / Reggiano G / Pike ACW / Wang D / McKinley G / Mukhopadhyay SMM / MacLean EM / Chalk R / Moreau C / Snee M / Bohstedt T / Singh NK / Abrusci P / Arrowsmith CH / Bountra C / Edwards AM / Marsden BD / Burgess-Brown NA / DiMaio F / Duerr KL | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Phospho-regulation, nucleotide binding and ion access control in potassium-chloride cotransporters. Authors: Gamma Chi / Rebecca Ebenhoch / Henry Man / Haiping Tang / Laurence E Tremblay / Gabriella Reggiano / Xingyu Qiu / Tina Bohstedt / Idlir Liko / Fernando G Almeida / Alexandre P Garneau / Dong ...Authors: Gamma Chi / Rebecca Ebenhoch / Henry Man / Haiping Tang / Laurence E Tremblay / Gabriella Reggiano / Xingyu Qiu / Tina Bohstedt / Idlir Liko / Fernando G Almeida / Alexandre P Garneau / Dong Wang / Gavin McKinley / Christophe P Moreau / Kiran D Bountra / Patrizia Abrusci / Shubhashish M M Mukhopadhyay / Alejandra Fernandez-Cid / Samira Slimani / Julie L Lavoie / Nicola A Burgess-Brown / Ben Tehan / Frank DiMaio / Ali Jazayeri / Paul Isenring / Carol V Robinson / Katharina L Dürr / ![]() ![]() ![]() Abstract: Potassium-coupled chloride transporters (KCCs) play crucial roles in regulating cell volume and intracellular chloride concentration. They are characteristically inhibited under isotonic conditions ...Potassium-coupled chloride transporters (KCCs) play crucial roles in regulating cell volume and intracellular chloride concentration. They are characteristically inhibited under isotonic conditions via phospho-regulatory sites located within the cytoplasmic termini. Decreased inhibitory phosphorylation in response to hypotonic cell swelling stimulates transport activity, and dysfunction of this regulatory process has been associated with various human diseases. Here, we present cryo-EM structures of human KCC3b and KCC1, revealing structural determinants for phospho-regulation in both N- and C-termini. We show that phospho-mimetic KCC3b is arrested in an inward-facing state in which intracellular ion access is blocked by extensive contacts with the N-terminus. In another mutant with increased isotonic transport activity, KCC1Δ19, this interdomain interaction is absent, likely due to a unique phospho-regulatory site in the KCC1 N-terminus. Furthermore, we map additional phosphorylation sites as well as a previously unknown ATP/ADP-binding pocket in the large C-terminal domain and show enhanced thermal stabilization of other CCCs by adenine nucleotides. These findings provide fundamentally new insights into the complex regulation of KCCs and may unlock innovative strategies for drug development. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 95.6 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 17.5 KB 17.5 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 10.8 KB | Display | ![]() |
Images | ![]() | 144.8 KB | ||
Masks | ![]() | 103 MB | ![]() | |
Others | ![]() ![]() | 95.9 MB 95.8 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6y5vMC ![]() 7ainC ![]() 7aioC ![]() 7aipC ![]() 7aiqC ![]() 7airC ![]() 7ngbC M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Map
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Voxel size | X=Y=Z: 0.83 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
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Density Histograms |
-Half map: #2
File | emd_10704_half_map_1.map | ||||||||||||
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Density Histograms |
-Half map: #1
File | emd_10704_half_map_2.map | ||||||||||||
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Density Histograms |
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Sample components
-Entire : Structure of Human Potassium Chloride Transporter KCC3b (S45D/T94...
Entire | Name: Structure of Human Potassium Chloride Transporter KCC3b (S45D/T940D/T997D) in KCl |
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Components |
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-Supramolecule #1: Structure of Human Potassium Chloride Transporter KCC3b (S45D/T94...
Supramolecule | Name: Structure of Human Potassium Chloride Transporter KCC3b (S45D/T940D/T997D) in KCl type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: ![]() |
Recombinant expression | Organism: ![]() |
Molecular weight | Theoretical: 240 KDa |
-Macromolecule #1: Solute carrier family 12 member 6
Macromolecule | Name: Solute carrier family 12 member 6 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 122.344422 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MPHFTVTKVE DPEEGAAASI SQEPSLADIK ARIQDSDEPD LSQNDITGEH SQLLDDGHKK ARNAYLNNSN YEEGDEYFDK NLALFEEEM DTRPKVSSLL NRMANYTNLT QGAKEHEEAE NITEGKKKPT KTPQMGTFMG VYLPCLQNIF GVILFLRLTW V VGTAGVLQ ...String: MPHFTVTKVE DPEEGAAASI SQEPSLADIK ARIQDSDEPD LSQNDITGEH SQLLDDGHKK ARNAYLNNSN YEEGDEYFDK NLALFEEEM DTRPKVSSLL NRMANYTNLT QGAKEHEEAE NITEGKKKPT KTPQMGTFMG VYLPCLQNIF GVILFLRLTW V VGTAGVLQ AFAIVLICCC CTMLTAISMS AIATNGVVPA GGSYFMISRA LGPEFGGAVG LCFYLGTTFA AAMYILGAIE IF LVYIVPR AAIFHSDDAL KESAAMLNNM RVYGTAFLVL MVLVVFIGVR YVNKFASLFL ACVIVSILAI YAGAIKSSFA PPH FPVCML GNRTLSSRHI DVCSKTKEIN NMTVPSKLWG FFCNSSQFFN ATCDEYFVHN NVTSIQGIPG LASGIITENL WSNY LPKGE IIEKPSAKSS DVLGSLNHEY VLVDITTSFT LLVGIFFPSV TGIMAGSNRS GDLKDAQKSI PIGTILAILT TSFVY LSNV VLFGACIEGV VLRDKFGDAV KGNLVVGTLS WPSPWVIVIG SFFSTCGAGL QSLTGAPRLL QAIAKDNIIP FLRVFG HSK ANGEPTWALL LTAAIAELGI LIASLDLVAP ILSMFFLMCY LFVNLACALQ TLLRTPNWRP RFRYYHWALS FMGMSIC LA LMFISSWYYA IVAMVIAGMI YKYIEYQGAE KEWGDGIRGL SLSAARFALL RLEEGPPHTK NWRPQLLVLL KLDEDLHV K HPRLLTFASQ LKAGKGLTIV GSVIVGNFLE NYGEALAAEQ TIKHLMEAEK VKGFCQLVVA AKLREGISHL IQSCGLGGM KHNTVVMGWP NGWRQSEDAR AWKTFIGTVR VTTAAHLALL VAKNISFFPS NVEQFSEGNI DVWWIVHDGG MLMLLPFLLK QHKVWRKCS IRIFTVAQLE DNSIQMKKDL ATFLYHLRIE AEVEVVEMHD SDISAYTYER DLMMEQRSQM LRHMRLSKTE R DREAQLVK DRNSMLRLTS IGSDEDEETE TYQEKVHMDW TKDKYMASRG QKAKSMEGFQ DLLNMRPDQS NVRRMHTAVK LN EVIVNKS HEAKLVLLNM PGPPRNPEGD ENYMEFLEVL TEGLERVLLV RGGGSEVITI YSA |
-Macromolecule #3: 2-acetamido-2-deoxy-beta-D-glucopyranose
Macromolecule | Name: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 3 / Number of copies: 2 / Formula: NAG |
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Molecular weight | Theoretical: 221.208 Da |
Chemical component information | ![]() ChemComp-NAG: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 6 mg/mL |
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Buffer | pH: 7.5 |
Grid | Model: Quantifoil, UltrAuFoil, R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY |
Vitrification | Cryogen name: ETHANE |
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Electron microscopy
Microscope | TFS KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |