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- PDB-7aio: Structure of Human Potassium Chloride Transporter KCC3 S45D/T940D... -
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Open data
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Basic information
Entry | Database: PDB / ID: 7aio | |||||||||
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Title | Structure of Human Potassium Chloride Transporter KCC3 S45D/T940D/T997D in NaCl (Subclass) | |||||||||
![]() | Solute carrier family 12 member 6 | |||||||||
![]() | TRANSPORT PROTEIN / CCC / transporter / human membrane protein / homodimer / KCC3 / KCC / potassium-chloride coupled transporter / Structural Genomics / Structural Genomics Consortium / SGC | |||||||||
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 / potassium ion transmembrane transport / chloride 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 | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.31 Å | |||||||||
![]() | Chi, G. / Man, H. / Ebenhoch, R. / Reggiano, G. / Pike, A.C.W. / Wang, D. / McKinley, G. / Mukhopadhyay, S.M.M. / MacLean, E.M. / Chalk, R. ...Chi, G. / Man, H. / Ebenhoch, R. / Reggiano, G. / Pike, A.C.W. / Wang, D. / McKinley, G. / Mukhopadhyay, S.M.M. / MacLean, E.M. / Chalk, R. / Moreau, C. / Snee, M. / Bohstedt, T. / Singh, N.K. / Abrusci, P. / Arrowsmith, C.H. / Bountra, C. / Edwards, A.M. / Marsden, B.D. / Burgess-Brown, N.A. / DiMaio, F. / Duerr, K.L. / Structural Genomics Consortium (SGC) | |||||||||
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
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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PDBx/mmCIF format | ![]() | 322.2 KB | Display | ![]() |
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PDB format | ![]() | 257.7 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 11800MC ![]() 6y5vC ![]() 7ainC ![]() 7aipC ![]() 7aiqC ![]() 7airC ![]() 7ngbC C: citing same article ( M: map data used to model this data |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 122273.344 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() #2: Polysaccharide | Source method: isolated from a genetically manipulated source #3: Sugar | Has ligand of interest | N | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Homodimeric complex of human potassium chloride transporter KCC3b Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT |
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Molecular weight | Value: 0.24 MDa / Experimental value: NO |
Source (natural) | Organism: ![]() |
Source (recombinant) | Organism: ![]() |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
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Processing
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Particle selection | Num. of particles selected: 4053596 | ||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.31 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 302166 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL Details: Reference model repositioned with Chimera and refined with Coot/Phenix. | ||||||||||||||||||||||||
Refinement | Cross valid method: NONE | ||||||||||||||||||||||||
Refine LS restraints |
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