[English] 日本語
Yorodumi
- EMDB-45081: Phosphorylated human NKCC1_K289NA492E in complex with furosemide -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-45081
TitlePhosphorylated human NKCC1_K289NA492E in complex with furosemide
Map dataPhosphorylated human NKCC1_K289NA492E in complex with furosemide
Sample
  • Complex: Phosphorylated NKCC1_K289NA492E in complex with furosemide
    • Protein or peptide: Solute carrier family 12 member 2
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: 5-(AMINOSULFONYL)-4-CHLORO-2-[(2-FURYLMETHYL)AMINO]BENZOIC ACID
  • Ligand: POTASSIUM ION
  • Ligand: CHLORIDE ION
  • Ligand: SODIUM ION
  • Ligand: MAGNESIUM ION
KeywordsSodium potassium chloride cotransporter / phosphorylation / outward-open state / furosemide / TRANSPORT PROTEIN
Function / homology
Function and homology information


positive regulation of cell volume / positive regulation of aspartate secretion / transepithelial ammonium transport / regulation of matrix metallopeptidase secretion / cell body membrane / metal ion transmembrane transporter activity / inorganic anion import across plasma membrane / inorganic cation import across plasma membrane / chloride:monoatomic cation symporter activity / sodium:potassium:chloride symporter activity ...positive regulation of cell volume / positive regulation of aspartate secretion / transepithelial ammonium transport / regulation of matrix metallopeptidase secretion / cell body membrane / metal ion transmembrane transporter activity / inorganic anion import across plasma membrane / inorganic cation import across plasma membrane / chloride:monoatomic cation symporter activity / sodium:potassium:chloride symporter activity / transepithelial chloride transport / Cation-coupled Chloride cotransporters / potassium ion transmembrane transporter activity / intracellular chloride ion homeostasis / negative regulation of vascular wound healing / ammonium transmembrane transport / sodium ion homeostasis / ammonium channel activity / chloride ion homeostasis / cell projection membrane / cellular response to potassium ion / T cell chemotaxis / cellular response to chemokine / potassium ion homeostasis / intracellular sodium ion homeostasis / hyperosmotic response / sodium ion import across plasma membrane / intracellular potassium ion homeostasis / cell volume homeostasis / regulation of spontaneous synaptic transmission / gamma-aminobutyric acid signaling pathway / maintenance of blood-brain barrier / potassium ion import across plasma membrane / lateral plasma membrane / transport across blood-brain barrier / monoatomic ion transport / sodium ion transmembrane transport / cytoplasmic vesicle membrane / chloride transmembrane transport / basal plasma membrane / cell periphery / cell projection / Hsp90 protein binding / extracellular vesicle / protein-folding chaperone binding / cell body / basolateral plasma membrane / neuron projection / apical plasma membrane / neuronal cell body / protein kinase binding / extracellular exosome / membrane / plasma membrane
Similarity search - Function
Solute carrier family 12 member 1/2 / Solute carrier family 12 member 2 / Amino acid permease, N-terminal / Amino acid permease N-terminal / SLC12A transporter, C-terminal / Solute carrier family 12 / Amino acid permease/ SLC12A domain / SLC12A transporter family / Amino acid permease
Similarity search - Domain/homology
Solute carrier family 12 member 2
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsZhao YX / Cao EH
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)5R01DK128592 United States
CitationJournal: EMBO J / Year: 2025
Title: Structural basis for human NKCC1 inhibition by loop diuretic drugs.
Authors: Yongxiang Zhao / Pietro Vidossich / Biff Forbush / Junfeng Ma / Jesse Rinehart / Marco De Vivo / Erhu Cao /
Abstract: Na-K-Cl cotransporters functions as an anion importers, regulating trans-epithelial chloride secretion, cell volume, and renal salt reabsorption. Loop diuretics, including furosemide, bumetanide, and ...Na-K-Cl cotransporters functions as an anion importers, regulating trans-epithelial chloride secretion, cell volume, and renal salt reabsorption. Loop diuretics, including furosemide, bumetanide, and torsemide, antagonize both NKCC1 and NKCC2, and are first-line medicines for the treatment of edema and hypertension. NKCC1 activation by the molecular crowding sensing WNK kinases is critical if cells are to combat shrinkage during hypertonic stress; however, how phosphorylation accelerates NKCC1 ion transport remains unclear. Here, we present co-structures of phospho-activated NKCC1 bound with furosemide, bumetanide, or torsemide showing that furosemide and bumetanide utilize a carboxyl group to coordinate and co-occlude a K, whereas torsemide encroaches and expels the K from the site. We also found that an amino-terminal segment of NKCC1, once phosphorylated, interacts with the carboxyl-terminal domain, and together, they engage with intracellular ion exit and appear to be poised to facilitate rapid ion translocation. Together, these findings enhance our understanding of NKCC-mediated epithelial ion transport and the molecular mechanisms of its inhibition by loop diuretics.
History
DepositionMay 25, 2024-
Header (metadata) releaseFeb 12, 2025-
Map releaseFeb 12, 2025-
UpdateMay 14, 2025-
Current statusMay 14, 2025Processing site: RCSB / Status: Released

-
Structure visualization

Supplemental images

emd_45081.png

Downloads & links

-
Map

FileDownload / File: emd_45081.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationPhosphorylated human NKCC1_K289NA492E in complex with furosemide
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
0.83 Å/pix.
x 360 pix.
= 297.216 Å		0.83 Å/pix.
x 360 pix.
= 297.216 Å		0.83 Å/pix.
x 360 pix.
= 297.216 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.8256 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.18
Minimum - Maximum-0.43376985 - 1.0456535
Average (Standard dev.)0.0005536064 (±0.024893336)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 297.216 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Additional map: Additional Map

Fileemd_45081_additional_1.map
AnnotationAdditional Map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Half Map B

Fileemd_45081_half_map_1.map
AnnotationHalf Map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Half Map A

Fileemd_45081_half_map_2.map
AnnotationHalf Map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : Phosphorylated NKCC1_K289NA492E in complex with furosemide

EntireName: Phosphorylated NKCC1_K289NA492E in complex with furosemide
Components
  • Complex: Phosphorylated NKCC1_K289NA492E in complex with furosemide
    • Protein or peptide: Solute carrier family 12 member 2
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: 5-(AMINOSULFONYL)-4-CHLORO-2-[(2-FURYLMETHYL)AMINO]BENZOIC ACID
  • Ligand: POTASSIUM ION
  • Ligand: CHLORIDE ION
  • Ligand: SODIUM ION
  • Ligand: MAGNESIUM ION

-
Supramolecule #1: Phosphorylated NKCC1_K289NA492E in complex with furosemide

SupramoleculeName: Phosphorylated NKCC1_K289NA492E in complex with furosemide
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

-
Macromolecule #1: Solute carrier family 12 member 2

MacromoleculeName: Solute carrier family 12 member 2 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 131.866328 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MEPRPTAPSS GAPGLAGVGE TPSAAALAAA RVELPGTAVP SVPEDAAPAS RDGGGVRDEG PAAAGDGLGR PLGPTPSQSR FQVDLVSEN AGRAAAAAAA AAAAAAAAGA GAGAKQTPAD GEASGESEPA KGSEEAKGRF RVNFVDPAAS SSAEDSLSDA A GVGVDGPN ...String:
MEPRPTAPSS GAPGLAGVGE TPSAAALAAA RVELPGTAVP SVPEDAAPAS RDGGGVRDEG PAAAGDGLGR PLGPTPSQSR FQVDLVSEN AGRAAAAAAA AAAAAAAAGA GAGAKQTPAD GEASGESEPA KGSEEAKGRF RVNFVDPAAS SSAEDSLSDA A GVGVDGPN VSFQNGGDTV LSEGSSLHSG GGGGSGHHQH YYYDTHTNTY YLR(TPO)FGHN(TPO)M DAVPRIDHYR H (TPO)AAQLGEK LLRPSLAELH DELEKEPFED GFANGEESTP TRDAVVTYTA ESKGVVKFGW INGVLVRCML NIWGVMLF I RLSWIVGQAG IGLSVLVIMM ATVVTTITGL STSAIATNGF VRGGGAYYLI SRSLGPEFGG AIGLIFAFAN AVAVAMYVV GFAETVVELL KEHSILMIDE INDIRIIGAI TVVILLGISV AGMEWEAKAQ IVLLVILLLA IGDFVIGTFI PLESKKPKGF FGYKSEIFN ENFGPDFREE ETFFSVFEIF FPAATGILAG ANISGDLADP QSAIPKGTLL AILITTLVYV GIAVSVGSCV V RDATGNVN DTIVTELTNC TSAACKLNFD FSSCESSPCS YGLMNNFQVM SMVSGFTPLI SAGIFSATLS SALASLVSAP KI FQALCKD NIYPAFQMFA KGYGKNNEPL RGYILTFLIA LGFILIAELN VIAPIISNFF LASYALINFS VFHASLAKSP GWR PAFKYY NMWISLLGAI LCCIVMFVIN WWAALLTYVI VLGLYIYVTY KKPDVNWGSS TQALTYLNAL QHSIRLSGVE DHVK NFRPQ CLVMTGAPNS RPALLHLVHD FTKNVGLMIC GHVHMGPRRQ AMKEMSIDQA KYQRWLIKNK MKAFYAPVHA DDLRE GAQY LMQAAGLGRM KPNTLVLGFK KDWLQADMRD VDMYINLFHD AFDIQYGVVV IRLKEGLDIS HLQGQEELLS SQEKSP GTK DVVVSVEYSK KSDLDTSKPL SEKPITHKVE EEDGKTATQP LLKKESKGPI VPLNVADQKL LEASTQFQKK QGKNTID VW WLFDDGGLTL LIPYLLTTKK KWKDCKIRVF IGGKINRIDH DRRAMATLLS KFRIDFSDIM VLGDINTKPK KENIIAFE E IIEPYRLHED DKEQDIADKM KEDEPWRITD NELELYKTKT YRQIRLNELL KEHSSTANII VMSLPVARKG AVSSALYMA WLEALSKDLP PILLVRGNHQ SVLTFYS

UniProtKB: Solute carrier family 12 member 2

-
Macromolecule #2: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 2 / Number of copies: 2 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

-
Macromolecule #3: 5-(AMINOSULFONYL)-4-CHLORO-2-[(2-FURYLMETHYL)AMINO]BENZOIC ACID

MacromoleculeName: 5-(AMINOSULFONYL)-4-CHLORO-2-[(2-FURYLMETHYL)AMINO]BENZOIC ACID
type: ligand / ID: 3 / Number of copies: 2 / Formula: FUN
Molecular weightTheoretical: 330.744 Da
Chemical component information

ChemComp-FUN:
5-(AMINOSULFONYL)-4-CHLORO-2-[(2-FURYLMETHYL)AMINO]BENZOIC ACID / medication*YM

-
Macromolecule #4: POTASSIUM ION

MacromoleculeName: POTASSIUM ION / type: ligand / ID: 4 / Number of copies: 2 / Formula: K
Molecular weightTheoretical: 39.098 Da

-
Macromolecule #5: CHLORIDE ION

MacromoleculeName: CHLORIDE ION / type: ligand / ID: 5 / Number of copies: 2 / Formula: CL
Molecular weightTheoretical: 35.453 Da

-
Macromolecule #6: SODIUM ION

MacromoleculeName: SODIUM ION / type: ligand / ID: 6 / Number of copies: 2
Molecular weightTheoretical: 22.99 Da

-
Macromolecule #7: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 7 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE

-
Electron microscopy

MicroscopeFEI MORGAGNI
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.5 µm / Nominal defocus min: 0.7000000000000001 µm

+
Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 8051818
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more