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- EMDB-24847: Locally refined protomer structure of native-form oocyte/egg Alph... -

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Entry
Database: EMDB / ID: EMD-24847
TitleLocally refined protomer structure of native-form oocyte/egg Alpha-2-Macroglobulin (A2Moo) tetramer
Map dataNative-form oocyte/egg Alpha-2-Macroglobulin (A2Moo) tetramer
Sample
  • Complex: nucleosome isolated from metaphase chromosome formed in Xenopus egg extract (oligo fraction)
    • Protein or peptide: Alpha 2-macroglobulin
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
KeywordsXenopus egg extract / Protease inhibitor / PROTEIN BINDING
Function / homology
Function and homology information


serine-type endopeptidase inhibitor activity / extracellular space
Similarity search - Function
Alpha-2-macroglobulin, TED domain / Alpha-2-macroglobulin, conserved site / Alpha-2-macroglobulin family thiolester region signature. / : / Alpha-macro-globulin thiol-ester bond-forming region / Alpha-macroglobulin, receptor-binding / Alpha-macroglobulin, receptor-binding domain superfamily / Macroglobulin domain MG4 / Macroglobulin domain MG3 / : ...Alpha-2-macroglobulin, TED domain / Alpha-2-macroglobulin, conserved site / Alpha-2-macroglobulin family thiolester region signature. / : / Alpha-macro-globulin thiol-ester bond-forming region / Alpha-macroglobulin, receptor-binding / Alpha-macroglobulin, receptor-binding domain superfamily / Macroglobulin domain MG4 / Macroglobulin domain MG3 / : / A-macroglobulin receptor binding domain / Macroglobulin domain MG4 / Macroglobulin domain MG3 / A-macroglobulin receptor / Alpha-2-macroglobulin / Macroglobulin domain / Alpha-2-macroglobulin, bait region domain / Alpha-macroglobulin-like, TED domain / Alpha-2-macroglobulin family / MG2 domain / A-macroglobulin TED domain / Alpha-2-macroglobulin bait region domain / Alpha-2-Macroglobulin / Alpha-2-macroglobulin family / Terpenoid cyclases/protein prenyltransferase alpha-alpha toroid / Immunoglobulin E-set / Immunoglobulin-like fold
Similarity search - Domain/homology
Uncharacterized protein LOC431886 isoform X1
Similarity search - Component
Biological speciesXenopus laevis (African clawed frog)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.65 Å
AuthorsArimura Y / Funabiki H
Funding support Japan, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM132111 Japan
Japan Society for the Promotion of Science (JSPS)JSPS Overseas Research Fellowships Japan
CitationJournal: J Mol Biol / Year: 2022
Title: Structural Mechanics of the Alpha-2-Macroglobulin Transformation.
Authors: Yasuhiro Arimura / Hironori Funabiki /
Abstract: Alpha-2-Macroglobulin (A2M) is the critical pan-protease inhibitor of the innate immune system. When proteases cleave the A2M bait region, global structural transformation of the A2M tetramer is ...Alpha-2-Macroglobulin (A2M) is the critical pan-protease inhibitor of the innate immune system. When proteases cleave the A2M bait region, global structural transformation of the A2M tetramer is triggered to entrap the protease. The structural basis behind the cleavage-induced transformation and the protease entrapment remains unclear. Here, we report cryo-EM structures of native- and intermediate-forms of the Xenopus laevis egg A2M homolog (A2Moo or ovomacroglobulin) tetramer at 3.7-4.1 Å and 6.4 Å resolution, respectively. In the native A2Moo tetramer, two pairs of dimers arrange into a cross-like configuration with four 60 Å-wide bait-exposing grooves. Each bait in the native form threads into an aperture formed by three macroglobulin domains (MG2, MG3, MG6). The bait is released from the narrowed aperture in the induced protomer of the intermediate form. We propose that the intact bait region works as a "latch-lock" to block futile A2M transformation until its protease-mediated cleavage.
History
DepositionSep 12, 2021-
Header (metadata) releaseJan 5, 2022-
Map releaseJan 5, 2022-
UpdateNov 13, 2024-
Current statusNov 13, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 8.75
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 8.75
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-7s62
  • Surface level: 8
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_24847.png

Downloads & links

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Map

FileDownload / File: emd_24847.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNative-form oocyte/egg Alpha-2-Macroglobulin (A2Moo) tetramer
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.47 Å/pix.
x 256 pix.
= 376.32 Å		1.47 Å/pix.
x 256 pix.
= 376.32 Å		1.47 Å/pix.
x 256 pix.
= 376.32 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.47 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 6.75 / Movie #1: 8.75
Minimum - Maximum-126.020200000000003 - 144.719510000000014
Average (Standard dev.)0.000000000006074 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 376.32 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.471.471.47
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z376.320376.320376.320
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ256256256
MAP C/R/S321
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-126.020144.7200.000

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Supplemental data

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Additional map: Native-form oocyte/egg Alpha-2-Macroglobulin (A2Moo) tetramer

Fileemd_24847_additional_1.map
AnnotationNative-form oocyte/egg Alpha-2-Macroglobulin (A2Moo) tetramer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Native-form oocyte/egg Alpha-2-Macroglobulin (A2Moo) tetramer

Fileemd_24847_half_map_1.map
AnnotationNative-form oocyte/egg Alpha-2-Macroglobulin (A2Moo) tetramer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Native-form oocyte/egg Alpha-2-Macroglobulin (A2Moo) tetramer

Fileemd_24847_half_map_2.map
AnnotationNative-form oocyte/egg Alpha-2-Macroglobulin (A2Moo) tetramer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : nucleosome isolated from metaphase chromosome formed in Xenopus e...

EntireName: nucleosome isolated from metaphase chromosome formed in Xenopus egg extract (oligo fraction)
Components
  • Complex: nucleosome isolated from metaphase chromosome formed in Xenopus egg extract (oligo fraction)
    • Protein or peptide: Alpha 2-macroglobulin
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: nucleosome isolated from metaphase chromosome formed in Xenopus e...

SupramoleculeName: nucleosome isolated from metaphase chromosome formed in Xenopus egg extract (oligo fraction)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Xenopus laevis (African clawed frog)

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Macromolecule #1: Alpha 2-macroglobulin

MacromoleculeName: Alpha 2-macroglobulin / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Molecular weightTheoretical: 159.480812 KDa
SequenceString: MSPNRFLLCV CILGLIAGGA AKVQYALTIP ALLKSGETQR ACVNLIGYHQ PLALSVVLEH QRVNISIFSE KVQPPHYFKC NKFMVPTVI TNAPDFVTLS VSGGGEDIKD RKAVVIAPLN TICLIQMDKP VYKPGCKVRF RLISLNTMLL PISEKYTAVY L EDPSGSRI ...String:
MSPNRFLLCV CILGLIAGGA AKVQYALTIP ALLKSGETQR ACVNLIGYHQ PLALSVVLEH QRVNISIFSE KVQPPHYFKC NKFMVPTVI TNAPDFVTLS VSGGGEDIKD RKAVVIAPLN TICLIQMDKP VYKPGCKVRF RLISLNTMLL PISEKYTAVY L EDPSGSRI AQWQNQESVG GVVQLEFPLI SDAAPGSYTI TAEGESCESA RQGFTVDEYI LPRFSVIVDP PNTISILDDI LT LNVSAIY TYGQPVPGSV TIKCCREASS YYGRKGNCFK GNRGICTNIT GELGPDGAFY GVVSLLPFQM GQSGFQMSLG VAL TVTEEG TGIQVTHQFF IMITSQLATL IFDYDALKEF YKRGIPYLVK VILTDANDNP MANEQVEVEL AGKTIGAVLT DKEG RAEYA IDTSSFVQEN FTVVVSYENP HQCYYTEWEG PDFPTAQHFV MRFYSETGSF LDIQGSSVEL NCGQVHNISV RYILS LDGM GEGATTATFY YLAMSRAKIV QHGQRDVHLN QSKSGLFNIG LNVTSDLAPG AELIVYCILD LELIADTISL DIEKCF QNQ VSLSFSDDLG PTASNVSLNL SAAPGSLCGV KVIDSSLLLI NPYESLSASG VYYSIPYLSL FGYNYGGFNL EEPEPPC ED PNTVIFCKGR YYLPVSSSTE GDTYQNLRRV GLVLGTSSKI RKPVVCGMEA KFSVPRKSSG ESDFGSSLSN GHVETLRK N FSETFLWRLV SVDSEGQNTI TETVPDTITK WQGSMFCVSE KEGFGITKYS ANFTSFLPFF VELSLPYSLT REEILVMKA FVSNYLEECI KIIVTLQPSA DFEVIPQDVK QDQCICSGGR SSYSWNIIAS SLGRISFIVS AETTHIGASC DGPSDQSQST RKDTVIQTI LVQPEGIRKE ETSSNLVCVE DSNVEMPINL TLPENIVQGS ASAFVTFVGD VLGLPLSNLQ NLLQMPYGCG E QNLARMAP IPYVLEYLNN TNQLTDELLQ TAVQFLNEGY YRQLRYKLPS GAYDAFWSSP SDGSSWLSAY TFKTFEKAKK YI YVDGKIQ QQTLLYLQTS QKLDNGCFKA EGNLFMRQCG QERDLCFTAY LAIALLESNY SSGMTLLDDA LGCLEAAMSS AST LYFKSY TVYVFTLVQN WEIRNTLLNE LKSKVVSERG TLHWEREDKL GQEGIPLYYP NYSPAEVEIT AYMLLSIAKG SDPT HDDLT YMAQISVWLI QQQNSYGGFR STQDTVVALQ ALAFYAQLLF KSNAHHNVFL RSEYGDVGQL NLSEHNRLVV QRLQL PEVS GNYSISINGT GCCLVQSTIR YNIPVPKENS AFYVAADSVS KNCLNGVAYT ITITVSVSYR GLRNETNMVI IDIQML SGY QADYPSLRQL ENSQQVSKTE EQDNHVFLYL NAVPLKTIQL SFKVLIGSRV LNVKSASVYV YDYYETGENG FASYSQP C

UniProtKB: Uncharacterized protein LOC431886 isoform X1

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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: 17 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

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

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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: 7.4
Component:
ConcentrationNameFormula
10.0 mMHEPES-KOH
30.0 mMKClKCl
1.0 mMEGTAEGTA
0.3 microg/mlLeupeptin
0.3 microg/mlPepstatin
0.3 microg/mlChymostatin
1.0 mMSodium Butyrate
1.0 mMbeta-glycerophosphate
1.0 %trehalose
0.1 %1,6,-hexanediol
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recording#0 - Image recording ID: 1 / #0 - Film or detector model: GATAN K2 SUMMIT (4k x 4k) / #0 - Detector mode: SUPER-RESOLUTION / #0 - Average electron dose: 33.11 e/Å2 / #1 - Image recording ID: 2 / #1 - Film or detector model: GATAN K2 SUMMIT (4k x 4k) / #1 - Detector mode: SUPER-RESOLUTION / #1 - Average electron dose: 38.34 e/Å2 / #2 - Image recording ID: 3 / #2 - Film or detector model: GATAN K2 SUMMIT (4k x 4k) / #2 - Detector mode: SUPER-RESOLUTION / #2 - Average electron dose: 35.27 e/Å2 / #3 - Image recording ID: 4 / #3 - Film or detector model: GATAN K2 SUMMIT (4k x 4k) / #3 - Detector mode: SUPER-RESOLUTION / #3 - Average electron dose: 34.2 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Image recording ID1
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.65 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.2.0) / Number images used: 195292
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 3.2.0)
FSC plot (resolution estimation)

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