[English] 日本語
Yorodumi
- EMDB-42072: Cryo-EM Structure of Brucella Abortus Lumazine Synthase (BLS) Eng... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-42072
TitleCryo-EM Structure of Brucella Abortus Lumazine Synthase (BLS) Engineered with Shiga Toxin I subunit B (Stx1B)
Map datatructure of Brucella Abortus Lumazine Synthase (BLS) Engineered with Shiga Toxin I subunit B (Stx1B)
Sample
  • Complex: Chimeric BLS-Stx1B protein
    • Protein or peptide: Shiga toxin subunit B,6,7-dimethyl-8-ribityllumazine synthase 2
KeywordsCHIMERA / SHIGA / TOXIN / IMMUNOGEN
Function / homology
Function and homology information


symbiont-mediated hemolysis of host erythrocyte / 6,7-dimethyl-8-ribityllumazine synthase / 6,7-dimethyl-8-ribityllumazine synthase activity / riboflavin synthase complex / riboflavin biosynthetic process / toxin activity / extracellular region / cytosol
Similarity search - Function
Shiga-like toxin, beta subunit / Shiga-like toxin beta subunit / Lumazine synthase / Lumazine/riboflavin synthase / Lumazine/riboflavin synthase superfamily / 6,7-dimethyl-8-ribityllumazine synthase / Enterotoxin
Similarity search - Domain/homology
6,7-dimethyl-8-ribityllumazine synthase 2 / Shiga toxin subunit B
Similarity search - Component
Biological speciesEscherichia coli O157:H7 (bacteria) / Brucella abortus biovar 1 (strain 9-941)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.37 Å
AuthorsCristofalo AE / Sharma A / Cerutti ML / Sharma K / Zylberman V / Goldbaum FA / Borgnia MJ / Otero LH
Funding support Argentina, United States, 2 items
OrganizationGrant numberCountry
Agencia Nacional de Promocion Cientifica y Tecnologica (FONCYT)PICT 2020-3047 Argentina
National Institutes of Health/National Institute of Environmental Health Sciences (NIH/NIEHS)ZIC ES103326 United States
CitationJournal: Protein Sci / Year: 2025
Title: Cryo-EM structures of engineered Shiga toxin-based immunogens capable of eliciting neutralizing antibodies with therapeutic potential against hemolytic uremic syndrome.
Authors: Alejandro Ezequiel Cristófalo / Arvind Sharma / María Laura Cerutti / Kedar Sharma / Roberto Melero / Romina Pardo / Fernando Alberto Goldbaum / Mario Borgnia / Vanesa Zylberman / Lisandro Horacio Otero /
Abstract: Shiga toxin-producing Escherichia coli-associated hemolytic uremic syndrome (STEC-HUS) is a serious disease that causes renal failure predominantly in children. Despite its significant impact, there ...Shiga toxin-producing Escherichia coli-associated hemolytic uremic syndrome (STEC-HUS) is a serious disease that causes renal failure predominantly in children. Despite its significant impact, there are currently no licensed vaccines or effective therapies available. The B subunits of Shiga toxins 1 and 2 (Stx1B and Stx2B) are suitable targets for developing neutralizing antibodies, but their pentameric assembly is unstable when isolated from the whole toxin. Taking advantage of the oligomeric symmetry shared between Stx1B and Stx2B with the lumazine synthase from Brucella spp. (BLS), we have previously engineered the chimeric toxoids BLS-Stx1B and BLS-Stx2B as immunogens to generate therapeutic equine polyclonal antibodies. The resulting product (INM004) has successfully passed Phases 1 and 2 clinical trials, and a Phase 3 has been launched in Argentina and seven European countries. In this work, we present the cryo-electron microscopy structures of BLS-Stx1B and BLS-Stx2B, which confirm that these engineered immunogens effectively stabilize the StxB pentamers. Moreover, our results reveal that both chimeric constructs present high flexibility at their extremes, corresponding to motions of the StxBs with respect to the BLS core. Additionally, we present structural evidence of the interaction between the chimeras and polyclonal Fab (pFab) fragments derived from INM004, demonstrating that the elicited neutralizing antibodies block most of the interaction surface of the toxins with their cellular receptors. These findings further validate this promising antibody-based therapy for mitigating STEC-HUS and demonstrate that the BLS-Stx1B and BLS-Stx2B chimeras are potential candidates for developing a human vaccine.
History
DepositionSep 22, 2023-
Header (metadata) releaseMar 26, 2025-
Map releaseMar 26, 2025-
UpdateOct 8, 2025-
Current statusOct 8, 2025Processing site: RCSB / Status: Released

-
Structure visualization

Supplemental images

emd_42072.png

Downloads & links

-
Map

FileDownload / File: emd_42072.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationtructure of Brucella Abortus Lumazine Synthase (BLS) Engineered with Shiga Toxin I subunit B (Stx1B)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
0.66 Å/pix.
x 480 pix.
= 316.8 Å		0.66 Å/pix.
x 480 pix.
= 316.8 Å		0.66 Å/pix.
x 480 pix.
= 316.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.66 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 2.4
Minimum - Maximum-12.529885 - 38.275481999999997
Average (Standard dev.)0.000000000532266 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions480480480
Spacing480480480
CellA=B=C: 316.80002 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Half map: Half Map 1

Fileemd_42072_half_map_1.map
AnnotationHalf Map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Half Map 2

Fileemd_42072_half_map_2.map
AnnotationHalf Map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : Chimeric BLS-Stx1B protein

EntireName: Chimeric BLS-Stx1B protein
Components
  • Complex: Chimeric BLS-Stx1B protein
    • Protein or peptide: Shiga toxin subunit B,6,7-dimethyl-8-ribityllumazine synthase 2

-
Supramolecule #1: Chimeric BLS-Stx1B protein

SupramoleculeName: Chimeric BLS-Stx1B protein / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Engineered chimera of Brucella abortus Lumazine Synthase (BLS) and Shiga Toxin 1 subunit B (Stx1B)
Source (natural)Organism: Escherichia coli O157:H7 (bacteria) / Location in cell: extracellular region

-
Macromolecule #1: Shiga toxin subunit B,6,7-dimethyl-8-ribityllumazine synthase 2

MacromoleculeName: Shiga toxin subunit B,6,7-dimethyl-8-ribityllumazine synthase 2
type: protein_or_peptide / ID: 1 / Number of copies: 10 / Enantiomer: LEVO / EC number: 6,7-dimethyl-8-ribityllumazine synthase
Source (natural)Organism: Brucella abortus biovar 1 (strain 9-941)
Molecular weightTheoretical: 25.39083 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MHTPDCVTGK VEYTKYNDDD TFTVKVGDKE LFTNRWNLQS LLLSAQITGM TVTIKTNACH NGGGFSEVIF RGSGSGSGSG SLKTSFKIA FIQARWHADI VDEARKSFVA ELAAKTGGSV EVEIFDVPGA YEIPLHAKTL ARTGRYAAIV GAAFVIDGGI Y RHDFVATA ...String:
MHTPDCVTGK VEYTKYNDDD TFTVKVGDKE LFTNRWNLQS LLLSAQITGM TVTIKTNACH NGGGFSEVIF RGSGSGSGSG SLKTSFKIA FIQARWHADI VDEARKSFVA ELAAKTGGSV EVEIFDVPGA YEIPLHAKTL ARTGRYAAIV GAAFVIDGGI Y RHDFVATA VINGMMQVQL ETEVPVLSVV LTPHHFHESK EHHDFFHAHF KVKGVEAAHA ALQIVSERSR IAALV

UniProtKB: Shiga toxin subunit B, 6,7-dimethyl-8-ribityllumazine synthase 2

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration1.4 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
137.0 mMNaClsodium chloride
2.7 mMKClpotassium chloride
10.0 mMNa2HPO4sodium hydrogen phosphate
1.8 mMKH2PO4potassium dihydrogen phosphate
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Support film - Film thickness: 50 / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE / Instrument: LEICA EM GP

-
Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
DetailsPreliminary grid screening was performed using SmartScope software
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number real images: 5884 / Average electron dose: 65.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 130000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

+
Image processing

Particle selectionNumber selected: 1773142
CTF correctionSoftware - Name: cryoSPARC (ver. 4.2.1) / Software - details: Patch CTF job was used / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE / Details: Ab-initio model was generated
Final reconstructionApplied symmetry - Point group: D5 (2x5 fold dihedral) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.37 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 88636
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1) / Software - details: Ab-initio job was used
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1) / Software - details: Homogeneous refinement job was used
Final 3D classificationSoftware - Name: cryoSPARC (ver. 4.2.1)

-
Atomic model buiding 1

Initial model
PDB IDChain

1xn1

source_name: PDB, initial_model_type: experimental model

1qnu

source_name: PDB, initial_model_type: experimental model
RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-8uav:
Cryo-EM Structure of Brucella Abortus Lumazine Synthase (BLS) Engineered with Shiga Toxin I subunit B (Stx1B)

+
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