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- PDB-7pqg: Structure of thermostabilised human NTCP in complex with nanobody 87 -

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Basic information

Entry
Database: PDB / ID: 7pqg
TitleStructure of thermostabilised human NTCP in complex with nanobody 87
Components
  • Nanobody 87
  • Sodium/bile acid cotransporter
KeywordsMEMBRANE PROTEIN / SLC10A1 / sodium bile acid symporter
Function / homology
Function and homology information


bile acid:sodium symporter activity / regulation of bile acid secretion / bile acid and bile salt transport / bile acid signaling pathway / Recycling of bile acids and salts / response to nutrient levels / response to organic cyclic compound / response to estrogen / cellular response to xenobiotic stimulus / virus receptor activity ...bile acid:sodium symporter activity / regulation of bile acid secretion / bile acid and bile salt transport / bile acid signaling pathway / Recycling of bile acids and salts / response to nutrient levels / response to organic cyclic compound / response to estrogen / cellular response to xenobiotic stimulus / virus receptor activity / basolateral plasma membrane / response to ethanol / plasma membrane
Similarity search - Function
Bile acid:sodium symporter/arsenical resistance protein Acr3 / Bile acid:sodium symporter / Sodium Bile acid symporter family / Sodium/solute symporter superfamily
Similarity search - Domain/homology
Hepatic sodium/bile acid cotransporter
Similarity search - Component
Biological speciesLama glama (llama)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsGoutam, K. / Reyes, N.
Funding supportEuropean Union, 1items
OrganizationGrant numberCountry
European Research Council (ERC)309657European Union
CitationJournal: Nature / Year: 2022
Title: Structural basis of sodium-dependent bile salt uptake into the liver.
Authors: Kapil Goutam / Francesco S Ielasi / Els Pardon / Jan Steyaert / Nicolas Reyes /
Abstract: The liver takes up bile salts from blood to generate bile, enabling absorption of lipophilic nutrients and excretion of metabolites and drugs. Human Na-taurocholate co-transporting polypeptide (NTCP) ...The liver takes up bile salts from blood to generate bile, enabling absorption of lipophilic nutrients and excretion of metabolites and drugs. Human Na-taurocholate co-transporting polypeptide (NTCP) is the main bile salt uptake system in liver. NTCP is also the cellular entry receptor of human hepatitis B and D viruses (HBV/HDV), and has emerged as an important target for antiviral drugs. However, the molecular mechanisms underlying NTCP transport and viral receptor functions remain incompletely understood. Here we present cryo-electron microscopy structures of human NTCP in complexes with nanobodies, revealing key conformations of its transport cycle. NTCP undergoes a conformational transition opening a wide transmembrane pore that serves as the transport pathway for bile salts, and exposes key determinant residues for HBV/HDV binding to the outside of the cell. A nanobody that stabilizes pore closure and inward-facing states impairs recognition of the HBV/HDV receptor-binding domain preS1, demonstrating binding selectivity of the viruses for open-to-outside over inward-facing conformations of the NTCP transport cycle. These results provide molecular insights into NTCP 'gated-pore' transport and HBV/HDV receptor recognition mechanisms, and are expected to help with development of liver disease therapies targeting NTCP.
History
DepositionSep 21, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0May 18, 2022Provider: repository / Type: Initial release
Revision 1.1Jul 6, 2022Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
B: Nanobody 87
A: Sodium/bile acid cotransporter


Theoretical massNumber of molelcules
Total (without water)51,6412
Polymers51,6412
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area1550 Å2
ΔGint-9 kcal/mol
Surface area20770 Å2
MethodPISA

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Components

#1: Antibody Nanobody 87


Mass: 14988.481 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lama glama (llama) / Strain: Lama glama / Variant: 9844 / Production host: Escherichia coli K-12 (bacteria) / Strain (production host): WK6
#2: Protein Sodium/bile acid cotransporter / Cell growth-inhibiting gene 29 protein / Na(+)/bile acid cotransporter / Na(+)/taurocholate ...Cell growth-inhibiting gene 29 protein / Na(+)/bile acid cotransporter / Na(+)/taurocholate transport protein / Sodium/taurocholate cotransporting polypeptide / Solute carrier family 10 member 1 / NTCP


Mass: 36652.391 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC10A1, NTCP, GIG29 / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: Q14973

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Human NTCP in complex with nanobody 87 / Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
11Lama glama (llama)9844
21Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
11Escherichia coli K-12 (bacteria)83333
21Homo sapiens (human)9606
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD
Image recordingElectron dose: 57.8 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.18.2_3874refinement
PHENIX1.18.2_3874refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 61053 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 156.22 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00413324
ELECTRON MICROSCOPYf_angle_d0.87674506
ELECTRON MICROSCOPYf_chiral_restr0.0493527
ELECTRON MICROSCOPYf_plane_restr0.0056551
ELECTRON MICROSCOPYf_dihedral_angle_d17.12391195

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