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- EMDB-48825: BSEP E297G Apo Structure with Disordered NBD2 in GDN -

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Basic information

Entry
Database: EMDB / ID: EMD-48825
TitleBSEP E297G Apo Structure with Disordered NBD2 in GDN
Map dataBSEP E297G Apo Structure with Disordered NBD2 in GDN
Sample
  • Cell: BSEP
KeywordsBSEP / ABCB11 / MEMBRANE PROTEIN
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.57 Å
AuthorsReddy BG / Gruget C / Moore J
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Commun Biol / Year: 2025
Title: A structural and mechanistic model for BSEP dysfunction in PFIC2 cholestatic disease.
Authors: Clémence Gruget / Bharat G Reddy / Jonathan M Moore /
Abstract: BSEP (ABCB11) transports bile salts across the canalicular membrane of hepatocytes, where they are incorporated into bile. Biallelic mutations in BSEP can cause Progressive Familial Intrahepatic ...BSEP (ABCB11) transports bile salts across the canalicular membrane of hepatocytes, where they are incorporated into bile. Biallelic mutations in BSEP can cause Progressive Familial Intrahepatic Cholestasis Type 2 (PFIC2), a rare pediatric disease characterized by hepatic bile acid accumulation leading to hepatotoxicity and, ultimately, liver failure. The most frequently occurring PFIC2 disease-causing mutations are missense mutations, which often display a phenotype with decreased protein expression and impaired maturation and trafficking to the canalicular membrane. To characterize the mutational effects on protein thermodynamic stability, we carried out biophysical characterization of 13 distinct PFIC2-associated variants using in-cell thermal shift (CETSA) measurements. These experiments reveal a cluster of residues localized to the NBD2-ICL2 interface, which exhibit severe destabilization relative to wild-type BSEP. A high-resolution (2.8 Å) cryo-EM structure provides a framework for rationalizing the CETSA results, revealing a novel, NBD2-localized mechanism through which the most severe missense patient mutations drive cholestatic disease. These findings suggest potential strategies for identifying mechanism-based small molecule correctors to address BSEP trafficking defects and advance novel therapies for PFIC2 and other cholestatic diseases.
History
DepositionJan 27, 2025-
Header (metadata) releaseApr 2, 2025-
Map releaseApr 2, 2025-
UpdateApr 23, 2025-
Current statusApr 23, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_48825.png

Downloads & links

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Map

FileDownload / File: emd_48825.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationBSEP E297G Apo Structure with Disordered NBD2 in GDN
Projections & slices

Image control

Size
Brightness
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AxesZ (Sec.)Y (Row.)X (Col.)
1.31 Å/pix.
x 256 pix.
= 334.848 Å		1.31 Å/pix.
x 256 pix.
= 334.848 Å		1.31 Å/pix.
x 256 pix.
= 334.848 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.308 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.12
Minimum - Maximum-0.13223808 - 0.2988893
Average (Standard dev.)-0.00023071765 (±0.01792383)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 334.848 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_48825_msk_1.map
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AxesZYX

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Half map: #1

Fileemd_48825_half_map_1.map
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Half map: #2

Fileemd_48825_half_map_2.map
Projections & Slices
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Sample components

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Entire : BSEP

EntireName: BSEP
Components
  • Cell: BSEP

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Supramolecule #1: BSEP

SupramoleculeName: BSEP / type: cell / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: In GDN Detergent
Source (natural)Organism: Homo sapiens (human)

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration2 mg/mL
BufferpH: 8
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Support film - Material: GOLD / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: EMDB MAP
EMDB ID:

31583

Final reconstructionResolution.type: BY AUTHOR / Resolution: 7.57 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 63241
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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Atomic model buiding 1

RefinementProtocol: FLEXIBLE FIT

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