[English] 日本語
Yorodumi
- PDB-8v8k: Crystal Structure of Nanobody NbE -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 8v8k
TitleCrystal Structure of Nanobody NbE
ComponentsNanobody NbE
KeywordsIMMUNE SYSTEM / Single Domain Antibody / Nanobody / VHH
Biological speciesLama glama (llama)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.85 Å
AuthorsKoehl, A. / Manglik, A. / Yu, J. / Kumar, A. / Zhang, X. / Martin, C. / Raia, P. / Steyaert, J. / Ballet, S. / Boland, A. / Stoeber, M.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute on Drug Abuse (NIH/NIDA)R61DA051531 United States
CitationJournal: Nat Commun / Year: 2024
Title: Structural basis of μ-opioid receptor targeting by a nanobody antagonist.
Authors: Jun Yu / Amit Kumar / Xuefeng Zhang / Charlotte Martin / Kevin Van Holsbeeck / Pierre Raia / Antoine Koehl / Toon Laeremans / Jan Steyaert / Aashish Manglik / Steven Ballet / Andreas Boland / Miriam Stoeber /
Abstract: The μ-opioid receptor (μOR), a prototypical G protein-coupled receptor (GPCR), is the target of opioid analgesics such as morphine and fentanyl. Due to the severe side effects of current opioid ...The μ-opioid receptor (μOR), a prototypical G protein-coupled receptor (GPCR), is the target of opioid analgesics such as morphine and fentanyl. Due to the severe side effects of current opioid drugs, there is considerable interest in developing novel modulators of μOR function. Most GPCR ligands today are small molecules, however biologics, including antibodies and nanobodies, represent alternative therapeutics with clear advantages such as affinity and target selectivity. Here, we describe the nanobody NbE, which selectively binds to the μOR and acts as an antagonist. We functionally characterize NbE as an extracellular and genetically encoded μOR ligand and uncover the molecular basis for μOR antagonism by determining the cryo-EM structure of the NbE-μOR complex. NbE displays a unique ligand binding mode and achieves μOR selectivity by interactions with the orthosteric pocket and extracellular receptor loops. Based on a β-hairpin loop formed by NbE that deeply protrudes into the μOR, we design linear and cyclic peptide analogs that recapitulate NbE's antagonism. The work illustrates the potential of nanobodies to uniquely engage with GPCRs and describes lower molecular weight μOR ligands that can serve as a basis for therapeutic developments.
History
DepositionDec 5, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 11, 2024Provider: repository / Type: Initial release
Revision 1.1Oct 2, 2024Group: Derived calculations / Category: pdbx_struct_assembly / pdbx_struct_assembly_gen
Revision 1.2Oct 23, 2024Group: Database references / Structure summary / Category: citation / citation_author / pdbx_entry_details
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

-
Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: Nanobody NbE
B: Nanobody NbE
C: Nanobody NbE


Theoretical massNumber of molelcules
Total (without water)40,0373
Polymers40,0373
Non-polymers00
Water00
1
A: Nanobody NbE


Theoretical massNumber of molelcules
Total (without water)13,3461
Polymers13,3461
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
B: Nanobody NbE


Theoretical massNumber of molelcules
Total (without water)13,3461
Polymers13,3461
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
C: Nanobody NbE


Theoretical massNumber of molelcules
Total (without water)13,3461
Polymers13,3461
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)76.559, 76.559, 298.596
Angle α, β, γ (deg.)90.00, 90.00, 120.00
Int Tables number181
Space group name H-MP6422

-
Components

#1: Antibody Nanobody NbE


Mass: 13345.769 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lama glama (llama) / Plasmid: Pet26b / Production host: Escherichia coli BL21(DE3) (bacteria)
Has protein modificationN

-
Experimental details

-
Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

-
Sample preparation

CrystalDensity Matthews: 3.15 Å3/Da / Density % sol: 61.01 % / Description: Hexagonal rods
Crystal growTemperature: 289 K / Method: vapor diffusion, hanging drop / pH: 7
Details: 0.1 M Hepes pH 7.0 1.0M Succinic Acid pH 7.0 1% PEG 2000 MME
PH range: 6.5-7.5 / Temp details: Incubator

-
Data collection

DiffractionMean temperature: 80 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 23-ID-D / Wavelength: 1.0332 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Aug 22, 2015
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.0332 Å / Relative weight: 1
ReflectionResolution: 2.85→49.58 Å / Num. obs: 12993 / % possible obs: 100 % / Redundancy: 11.9 % / CC1/2: 0.996 / Net I/σ(I): 12.3
Reflection shellResolution: 2.85→2.95 Å / Mean I/σ(I) obs: 1.2 / Num. unique obs: 1258 / CC1/2: 0.538

-
Processing

Software
NameVersionClassification
PHENIX(1.19.2_4158: ???)refinement
Aimlessdata scaling
XDSdata reduction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.85→44.37 Å / SU ML: 0.52 / Cross valid method: FREE R-VALUE / σ(F): 0 / Phase error: 35.32 / Stereochemistry target values: ML
Details: Iterative manual rebuilding in coot with automated refinement in Phenix
RfactorNum. reflection% reflectionSelection details
Rfree0.3313 1288 10.04 %Random
Rwork0.286 ---
obs0.2906 12826 98.71 %-
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 2.85→44.37 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2701 0 0 0 2701
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0022758
X-RAY DIFFRACTIONf_angle_d0.5753730
X-RAY DIFFRACTIONf_dihedral_angle_d11.338963
X-RAY DIFFRACTIONf_chiral_restr0.041397
X-RAY DIFFRACTIONf_plane_restr0.003473
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.85-2.960.50361340.43061185X-RAY DIFFRACTION95
2.96-3.10.42251350.36231205X-RAY DIFFRACTION97
3.1-3.260.35611380.33521229X-RAY DIFFRACTION98
3.26-3.470.36711420.29211265X-RAY DIFFRACTION99
3.47-3.730.36291400.28671266X-RAY DIFFRACTION100
3.73-4.110.32421410.28491283X-RAY DIFFRACTION100
4.11-4.70.30541470.24641307X-RAY DIFFRACTION100
4.7-5.920.28711480.25541339X-RAY DIFFRACTION100
5.92-100.32471630.29621459X-RAY DIFFRACTION100

+
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