[English] 日本語
Yorodumi
- EMDB-6189: Negative stain single particle electron microscopy of unliganded ... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-6189
TitleNegative stain single particle electron microscopy of unliganded 16055 SOSIP
Map dataReconstruction of unliganded 16055 clade C SOSIP
Sample
  • Sample: 16055 SOSIP clade C HIV, unliganded
  • Protein or peptide: 16055 SOSIP gp140
Biological speciesSimian-Human immunodeficiency virus
Methodsingle particle reconstruction / negative staining / Resolution: 18.0 Å
Authorsde Val N / Guenaga J / Ward AB / Wyatt RT
CitationJournal: PLoS Pathog / Year: 2015
Title: Well-ordered trimeric HIV-1 subtype B and C soluble spike mimetics generated by negative selection display native-like properties.
Authors: Javier Guenaga / Natalia de Val / Karen Tran / Yu Feng / Karen Satchwell / Andrew B Ward / Richard T Wyatt /
Abstract: The structure of BG505 gp140 SOSIP, a soluble mimic of the native HIV-1 envelope glycoprotein (Env), marks the beginning of new era in Env structure-based immunogen design. Displaying a well-ordered ...The structure of BG505 gp140 SOSIP, a soluble mimic of the native HIV-1 envelope glycoprotein (Env), marks the beginning of new era in Env structure-based immunogen design. Displaying a well-ordered quaternary structure, these subtype A-derived trimers display an excellent antigenic profile, discriminating recognition by broadly neutralizing antibodies (bNAbs) from non-broadly neutralizing antibodies (non-bNAbs), and provide a solid Env-based immunogenic platform starting point. Even with this important advance, obtaining homogeneous well-ordered soluble SOSIP trimers derived from other subtypes remains challenging. Here, we report the "rescue" of homogeneous well-ordered subtype B and C SOSIP trimers from a heterogeneous Env mixture using CD4 binding site-directed (CD4bs) non-bNAbs in a negative-selection purification process. These non-bNAbs recognize the primary receptor CD4bs only on disordered trimers but not on the native Env spike or well-ordered soluble trimers due to steric hindrance. Following negative selection to remove disordered oligomers, we demonstrated recovery of well-ordered, homogeneous trimers by electron microscopy (EM). We obtained 3D EM reconstructions of unliganded trimers, as well as in complex with sCD4, a panel of CD4bs-directed bNAbs, and the cleavage-dependent, trimer-specific bNAb, PGT151. Using bio-layer light interferometry (BLI) we demonstrated that the well-ordered trimers were efficiently recognized by bNAbs and poorly recognized by non-bNAbs, representing soluble mimics of the native viral spike. Biophysical characterization was consistent with the thermostability of a homogeneous species that could be further stabilized by specific bNAbs. This study revealed that Env trimers generate different frequencies of well-ordered versus disordered aberrant trimers even when they are genetically identical. By negatively selecting the native-like well-ordered trimers, we establish a new means to obtain soluble Env mimetics derived from subtypes B and C for expanded use as candidate vaccine immunogens.
History
DepositionNov 19, 2014-
Header (metadata) releaseDec 24, 2014-
Map releaseJan 21, 2015-
UpdateJan 21, 2015-
Current statusJan 21, 2015Processing site: RCSB / Status: Released

-
Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 17.1
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 17.1
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

-
Map

FileDownload / File: emd_6189.map.gz / Format: CCP4 / Size: 15.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of unliganded 16055 clade C SOSIP
Voxel sizeX=Y=Z: 2.05 Å
Density
Contour LevelBy EMDB: 17.100000000000001 / Movie #1: 17.1
Minimum - Maximum-13.41003227 - 66.031448359999999
Average (Standard dev.)0.50939649 (±3.49403405)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions160160160
Spacing160160160
CellA=B=C: 328.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.052.052.05
M x/y/z160160160
origin x/y/z0.0000.0000.000
length x/y/z328.000328.000328.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-72-72-72
NX/NY/NZ145145145
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS160160160
D min/max/mean-13.41066.0310.509

-
Supplemental data

-
Sample components

-
Entire : 16055 SOSIP clade C HIV, unliganded

EntireName: 16055 SOSIP clade C HIV, unliganded
Components
  • Sample: 16055 SOSIP clade C HIV, unliganded
  • Protein or peptide: 16055 SOSIP gp140

-
Supramolecule #1000: 16055 SOSIP clade C HIV, unliganded

SupramoleculeName: 16055 SOSIP clade C HIV, unliganded / type: sample / ID: 1000 / Oligomeric state: trimer / Number unique components: 1
Molecular weightExperimental: 420 KDa / Theoretical: 420 KDa / Method: Size exclusion chromatography (SEC)

-
Macromolecule #1: 16055 SOSIP gp140

MacromoleculeName: 16055 SOSIP gp140 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Oligomeric state: trimer / Recombinant expression: Yes
Source (natural)Organism: Simian-Human immunodeficiency virus
Molecular weightExperimental: 420 KDa / Theoretical: 420 KDa
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK 293T

-
Experimental details

-
Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration0.5 mg/mL
BufferpH: 7.4 / Details: 50 mM Tris-HCl, 150 mM NaCl
StainingType: NEGATIVE
Details: Grids were stained for 30 seconds with 2% uranyl formate.
GridDetails: 400 Cu mesh grids, glow-discharged at 20 mA for 30 seconds
VitrificationCryogen name: NONE / Instrument: OTHER

-
Electron microscopy

MicroscopeFEI TECNAI SPIRIT
Electron beamAcceleration voltage: 120 kV / Electron source: LAB6
Electron opticsCalibrated magnification: 52000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1.0 µm / Nominal defocus min: 0.75 µm / Nominal magnification: 46000
Specialist opticsEnergy filter - Name: FEI
Sample stageSpecimen holder model: OTHER / Tilt angle max: 50
DateNov 22, 2013
Image recordingCategory: CCD / Film or detector model: DIRECT ELECTRON DE-10 (5k x 4k) / Number real images: 266 / Average electron dose: 35.45 e/Å2
Tilt angle min0
Experimental equipment
Model: Tecnai Spirit / Image courtesy: FEI Company

-
Image processing

CTF correctionDetails: CTFFindV3
Final reconstructionResolution.type: BY AUTHOR / Resolution: 18.0 Å / Resolution method: OTHER / Software - Name: EMAN2, sparx / Number images used: 53573

+
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