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- EMDB-20616: EM structure of MPEG-1(w.t.) soluble pre-pore -

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Basic information

Entry
Database: EMDB / ID: EMD-20616
TitleEM structure of MPEG-1(w.t.) soluble pre-pore
Map data
Sample
  • Complex: Macrophage-Expressed Gene 1 (MPEG-1) 16-mer soluble pre-pore complex
    • Protein or peptide: Macrophage-expressed gene 1 protein
KeywordsMPEG-1/Perforin-2 / MACPF / macrophage / pore forming proteins / immunity / IMMUNE SYSTEM
Function / homology
Function and homology information


dendritic cell antigen processing and presentation / antigen processing and presentation of exogenous peptide antigen / phagolysosome membrane / antibacterial innate immune response / wide pore channel activity / antigen processing and presentation of exogenous peptide antigen via MHC class I / phagocytic vesicle / phagocytic vesicle membrane / cytoplasmic vesicle / defense response to Gram-negative bacterium ...dendritic cell antigen processing and presentation / antigen processing and presentation of exogenous peptide antigen / phagolysosome membrane / antibacterial innate immune response / wide pore channel activity / antigen processing and presentation of exogenous peptide antigen via MHC class I / phagocytic vesicle / phagocytic vesicle membrane / cytoplasmic vesicle / defense response to Gram-negative bacterium / adaptive immune response / defense response to Gram-positive bacterium / defense response to bacterium / extracellular region
Similarity search - Function
Macrophage-expressed gene 1 protein / membrane-attack complex / perforin / MAC/Perforin domain / Membrane attack complex/perforin (MACPF) domain profile. / Membrane attack complex component/perforin (MACPF) domain
Similarity search - Domain/homology
Macrophage-expressed gene 1 protein
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.49 Å
AuthorsPang SS / Bayly-Jones C
Funding support Australia, 1 items
OrganizationGrant numberCountry
Australian Research Council (ARC) Australia
CitationJournal: Nat Commun / Year: 2019
Title: The cryo-EM structure of the acid activatable pore-forming immune effector Macrophage-expressed gene 1.
Authors: Siew Siew Pang / Charles Bayly-Jones / Mazdak Radjainia / Bradley A Spicer / Ruby H P Law / Adrian W Hodel / Edward S Parsons / Susan M Ekkel / Paul J Conroy / Georg Ramm / Hariprasad ...Authors: Siew Siew Pang / Charles Bayly-Jones / Mazdak Radjainia / Bradley A Spicer / Ruby H P Law / Adrian W Hodel / Edward S Parsons / Susan M Ekkel / Paul J Conroy / Georg Ramm / Hariprasad Venugopal / Phillip I Bird / Bart W Hoogenboom / Ilia Voskoboinik / Yann Gambin / Emma Sierecki / Michelle A Dunstone / James C Whisstock /
Abstract: Macrophage-expressed gene 1 (MPEG1/Perforin-2) is a perforin-like protein that functions within the phagolysosome to damage engulfed microbes. MPEG1 is thought to form pores in target membranes, ...Macrophage-expressed gene 1 (MPEG1/Perforin-2) is a perforin-like protein that functions within the phagolysosome to damage engulfed microbes. MPEG1 is thought to form pores in target membranes, however, its mode of action remains unknown. We use cryo-Electron Microscopy (cryo-EM) to determine the 2.4 Å structure of a hexadecameric assembly of MPEG1 that displays the expected features of a soluble prepore complex. We further discover that MPEG1 prepore-like assemblies can be induced to perforate membranes through acidification, such as would occur within maturing phagolysosomes. We next solve the 3.6 Å cryo-EM structure of MPEG1 in complex with liposomes. These data reveal that a multi-vesicular body of 12 kDa (MVB12)-associated β-prism (MABP) domain binds membranes such that the pore-forming machinery of MPEG1 is oriented away from the bound membrane. This unexpected mechanism of membrane interaction suggests that MPEG1 remains bound to the phagolysosome membrane while simultaneously forming pores in engulfed bacterial targets.
History
DepositionAug 19, 2019-
Header (metadata) releaseSep 25, 2019-
Map releaseSep 25, 2019-
UpdateOct 30, 2024-
Current statusOct 30, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 1
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6u23
  • Surface level: 1
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20616.png

Downloads & links

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Map

FileDownload / File: emd_20616.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 400 pix.
= 440. Å		1.1 Å/pix.
x 400 pix.
= 440. Å		1.1 Å/pix.
x 400 pix.
= 440. Å

Surface

Projections

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.0 / Movie #1: 1
Minimum - Maximum-3.6582363 - 6.978327
Average (Standard dev.)-0.0005539284 (±0.18564408)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 440.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.11.11.1
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z440.000440.000440.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ320320320
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-3.6586.978-0.001

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Supplemental data

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Mask #1

Fileemd_20616_msk_1.map
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Additional map: unsharpened map

Fileemd_20616_additional.map
Annotationunsharpened map
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Half map: #2

Fileemd_20616_half_map_1.map
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Half map: #1

Fileemd_20616_half_map_2.map
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Sample components

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Entire : Macrophage-Expressed Gene 1 (MPEG-1) 16-mer soluble pre-pore complex

EntireName: Macrophage-Expressed Gene 1 (MPEG-1) 16-mer soluble pre-pore complex
Components
  • Complex: Macrophage-Expressed Gene 1 (MPEG-1) 16-mer soluble pre-pore complex
    • Protein or peptide: Macrophage-expressed gene 1 protein

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Supramolecule #1: Macrophage-Expressed Gene 1 (MPEG-1) 16-mer soluble pre-pore complex

SupramoleculeName: Macrophage-Expressed Gene 1 (MPEG-1) 16-mer soluble pre-pore complex
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 2.3 MDa

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Macromolecule #1: Macrophage-expressed gene 1 protein

MacromoleculeName: Macrophage-expressed gene 1 protein / type: protein_or_peptide / ID: 1 / Number of copies: 16 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 71.110125 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: KSGKPSGEMD EVGVQKCKNA LKLPVLEVLP GGGWDNLRNV DMGRVMELTY SNCRTTEDGQ YIIPDEIFTI PQKQSNLEMN SEILESWAN YQSSTSYSIN TELSLFSKVN GKFSTEFQRM KTLQVKDQAI TTRVQVRNLV YTVKINPTLE LSSGFRKELL D ISDRLENN ...String:
KSGKPSGEMD EVGVQKCKNA LKLPVLEVLP GGGWDNLRNV DMGRVMELTY SNCRTTEDGQ YIIPDEIFTI PQKQSNLEMN SEILESWAN YQSSTSYSIN TELSLFSKVN GKFSTEFQRM KTLQVKDQAI TTRVQVRNLV YTVKINPTLE LSSGFRKELL D ISDRLENN QTRMATYLAE LLVLNYGTHV TTSVDAGAAL IQEDHLRASF LQDSQSSRSA VTASAGLAFQ NTVNFKFEEN YT SQNVLTK SYLSNRTNSR VQSIGGVPFY PGITLQAWQQ GITNHLVAID RSGLPLHFFI NPNMLPDLPG PLVKKVSKTV ETA VKRYYT FNTYPGCTDL NSPNFNFQAN TDDGSCEGKM TNFSFGGVYQ ECTQLSGNRD VLLCQKLEQK NPLTGDFSCP SGYS PVHLL SQIHEEGYNH LECHRKCTLL VFCKTVCEDV FQVAKAEFRA FWCVASSQVP ENSGLLFGGL FSSKSINPMT NAQSC PAGY FPLRLFENLK VCVSQDYELG SRFAVPFGGF FSCTVGNPLV DPAISRDLGA PSLKKCPGGF SQHPALISDG CQVSYC VKS GLFTGGSLPP ARLPPFTRPP LMSQAATNTV IVTNSENARS WIKDSQTHQW RLGEPIELRR AMNVIHGDGG GLSHHHH HH

UniProtKB: Macrophage-expressed gene 1 protein

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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: 7.2
Component:
ConcentrationFormulaName
20.0 mMTris-HClTris buffer
0.3 MNaClsalt
GridModel: Quantifoil R1.2/1.3 / Material: COPPER
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
DetailsThis sample was monodisperse

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Average electron dose: 45.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 202326
Startup modelType of model: OTHER
Final reconstructionApplied symmetry - Point group: C16 (16 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.49 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1) / Number images used: 25359
Initial angle assignmentType: COMMON LINE
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: AB INITIO MODEL
Output model

PDB-6u23:
EM structure of MPEG-1(w.t.) soluble pre-pore

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