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- EMDB-15072: Structure of murine perforin-2 (Mpeg1) pore in ring form -

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Basic information

Entry
Database: EMDB / ID: EMD-15072
TitleStructure of murine perforin-2 (Mpeg1) pore in ring form
Map dataFinal map, local resolution filtered
Sample
  • Complex: murine perforin-2 ectodomain in flat ring pore form
    • Protein or peptide: Macrophage-expressed gene 1 protein
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: CYCLOHEXYL-HEXYL-BETA-D-MALTOSIDE
Function / homology
Function and homology information


dendritic cell antigen processing and presentation / antigen processing and presentation of exogenous peptide antigen / phagolysosome membrane / endolysosome / pore-forming activity / 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 ...dendritic cell antigen processing and presentation / antigen processing and presentation of exogenous peptide antigen / phagolysosome membrane / endolysosome / pore-forming activity / 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
Similarity search - Function
Macrophage-expressed gene 1 protein / membrane-attack complex / perforin / Membrane attack complex/perforin (MACPF) domain profile. / MAC/Perforin domain / Membrane attack complex component/perforin (MACPF) domain
Similarity search - Domain/homology
Macrophage-expressed gene 1 protein
Similarity search - Component
Biological speciesMus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.0 Å
AuthorsYu X / Ni T / Zhang P / Gilbert R
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Wellcome Trust United Kingdom
CitationJournal: EMBO J / Year: 2022
Title: Cryo-EM structures of perforin-2 in isolation and assembled on a membrane suggest a mechanism for pore formation.
Authors: Xiulian Yu / Tao Ni / George Munson / Peijun Zhang / Robert J C Gilbert /
Abstract: Perforin-2 (PFN2, MPEG1) is a key pore-forming protein in mammalian innate immunity restricting intracellular bacteria proliferation. It forms a membrane-bound pre-pore complex that converts to a ...Perforin-2 (PFN2, MPEG1) is a key pore-forming protein in mammalian innate immunity restricting intracellular bacteria proliferation. It forms a membrane-bound pre-pore complex that converts to a pore-forming structure upon acidification; but its mechanism of conformational transition has been debated. Here we used cryo-electron microscopy, tomography and subtomogram averaging to determine structures of PFN2 in pre-pore and pore conformations in isolation and bound to liposomes. In isolation and upon acidification, the pre-assembled complete pre-pore rings convert to pores in both flat ring and twisted conformations. On membranes, in situ assembled PFN2 pre-pores display various degrees of completeness; whereas PFN2 pores are mainly incomplete arc structures that follow the same subunit packing arrangements as found in isolation. Both assemblies on membranes use their P2 β-hairpin for binding to the lipid membrane surface. Overall, these structural snapshots suggest a molecular mechanism for PFN2 pre-pore to pore transition on a targeted membrane, potentially using the twisted pore as an intermediate or alternative state to the flat conformation, with the capacity to cause bilayer distortion during membrane insertion.
History
DepositionJun 1, 2022-
Header (metadata) releaseJul 20, 2022-
Map releaseJul 20, 2022-
UpdateDec 14, 2022-
Current statusDec 14, 2022Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15072.png

Downloads & links

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Map

FileDownload / File: emd_15072.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFinal map, local resolution filtered
Voxel sizeX=Y=Z: 1.047 Å
Density
Contour LevelBy AUTHOR: 0.0225
Minimum - Maximum-0.17784402 - 0.3156616
Average (Standard dev.)0.00015892243 (±0.0071583525)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 376.92 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_15072_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map2 from relion

Fileemd_15072_half_map_1.map
Annotationhalf map2 from relion
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map1 from relion

Fileemd_15072_half_map_2.map
Annotationhalf map1 from relion
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : murine perforin-2 ectodomain in flat ring pore form

EntireName: murine perforin-2 ectodomain in flat ring pore form
Components
  • Complex: murine perforin-2 ectodomain in flat ring pore form
    • Protein or peptide: Macrophage-expressed gene 1 protein
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: CYCLOHEXYL-HEXYL-BETA-D-MALTOSIDE

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Supramolecule #1: murine perforin-2 ectodomain in flat ring pore form

SupramoleculeName: murine perforin-2 ectodomain in flat ring pore form / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Mus musculus (house mouse)

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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: Mus musculus (house mouse)
Molecular weightTheoretical: 71.359812 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: ETGDKPLGET GTTGFQICKN ALKLPVLEVL PGGGWDNLRN VDMGRVMDLT YTNCKTTEDG QYIIPDEVYT IPQKESNLEM NSEVLESWM NYQSTTSLSI NTELALFSRV NGKFSTEFQR MKTLQVKDQA VTTRVQVRNR IYTVKTTPTS ELSLGFTKAL M DICDQLEK ...String:
ETGDKPLGET GTTGFQICKN ALKLPVLEVL PGGGWDNLRN VDMGRVMDLT YTNCKTTEDG QYIIPDEVYT IPQKESNLEM NSEVLESWM NYQSTTSLSI NTELALFSRV NGKFSTEFQR MKTLQVKDQA VTTRVQVRNR IYTVKTTPTS ELSLGFTKAL M DICDQLEK NQTKMATYLA ELLILNYGTH VITSVDAGAA LVQEDHVRSS FLLDNQNSQN TVTASAGIAF LNIVNFKVET DY ISQTSLT KDYLSNRTNS RVQSFGGVPF YPGITLETWQ KGITNHLVAI DRAGLPLHFF IKPDKLPGLP GPLVKKLSKT VET AVRHYY TFNTHPGCTN VDSPNFNFQA NMDDDSCDAK VTNFTFGGVY QECTELSGDV LCQNLEQKNL LTGDFSCPPG YSPV HLLSQ THEEGYSRLE CKKKCTLKIF CKTVCEDVFR VAKAEFRAYW CVAAGQVPDN SGLLFGGVFT DKTINPMTNA QSCPA GYIP LNLFESLKVC VSLDYELGFK FSVPFGGFFS CIMGNPLVNS DTAKDVRAPS LKKCPGGFSQ HLAVISDGCQ VSYCVK AGI FTGGSLLPVR LPPYTKPPLM SQVATNTVIV TNSETARSWI KDPQTNQWKL GEPLELRRAM TVIHGDSNGM SGGGTKT ET SQVAPA

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Macromolecule #2: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 2 / Number of copies: 16 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #3: CYCLOHEXYL-HEXYL-BETA-D-MALTOSIDE

MacromoleculeName: CYCLOHEXYL-HEXYL-BETA-D-MALTOSIDE / type: ligand / ID: 3 / Number of copies: 16 / Formula: MA4
Molecular weightTheoretical: 508.6 Da
Chemical component information

ChemComp-MA4:
CYCLOHEXYL-HEXYL-BETA-D-MALTOSIDE

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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

BufferpH: 3.8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.4 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:

10135

Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 519614
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-8a1d:
Structure of murine perforin-2 (Mpeg1) pore in ring form

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