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- EMDB-15220: Partial dimer complex of PAPP-A and its inhibitor STC2 -

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Basic information

Entry
Database: EMDB / ID: EMD-15220
TitlePartial dimer complex of PAPP-A and its inhibitor STC2
Map dataPrimary, sharpened map of a partial dimer of PAPP-A in complex with a STC2 subunit
Sample
  • Complex: Partial PAPP-A dimer in complex with subunit of its endogenous inhibitor STC2 dimer
    • Complex: PAPP-A
      • Complex: N-terminal part of PAPP-A
        • Protein or peptide: Pappalysin-1
      • Complex: C-terminal part of PAPP-A
    • Complex: Subunit of Stanniocalcin-2
      • Protein or peptide: Stanniocalcin-2
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: ZINC ION
  • Ligand: CALCIUM IONCalcium
Function / homology
Function and homology information


regulation of hormone biosynthetic process / pappalysin-1 / response to follicle-stimulating hormone / regulation of store-operated calcium entry / response to vitamin D / protein metabolic process / negative regulation of multicellular organism growth / response to dexamethasone / decidualization / endoplasmic reticulum unfolded protein response ...regulation of hormone biosynthetic process / pappalysin-1 / response to follicle-stimulating hormone / regulation of store-operated calcium entry / response to vitamin D / protein metabolic process / negative regulation of multicellular organism growth / response to dexamethasone / decidualization / endoplasmic reticulum unfolded protein response / embryo implantation / female pregnancy / Post-translational protein phosphorylation / protein catabolic process / hormone activity / metalloendopeptidase activity / response to peptide hormone / intracellular calcium ion homeostasis / metallopeptidase activity / Regulation of Insulin-like Growth Factor (IGF) transport and uptake by Insulin-like Growth Factor Binding Proteins (IGFBPs) / cellular response to hypoxia / response to oxidative stress / cell surface receptor signaling pathway / endoplasmic reticulum lumen / negative regulation of gene expression / heme binding / perinuclear region of cytoplasm / Golgi apparatus / enzyme binding / endoplasmic reticulum / protein homodimerization activity / proteolysis / extracellular space / zinc ion binding / extracellular region
Similarity search - Function
Stanniocalcin / Stanniocalcin family / Pappalysin-1/2 / Myxococcus cysteine-rich repeat / LamG-like jellyroll fold / LamG-like jellyroll fold domain / Peptidase M43, pregnancy-associated plasma-A / Pregnancy-associated plasma protein-A / Concanavalin A-like lectin/glucanases superfamily / Notch domain ...Stanniocalcin / Stanniocalcin family / Pappalysin-1/2 / Myxococcus cysteine-rich repeat / LamG-like jellyroll fold / LamG-like jellyroll fold domain / Peptidase M43, pregnancy-associated plasma-A / Pregnancy-associated plasma protein-A / Concanavalin A-like lectin/glucanases superfamily / Notch domain / Domain found in Notch and Lin-12 / Sushi repeat (SCR repeat) / Domain abundant in complement control proteins; SUSHI repeat; short complement-like repeat (SCR) / Sushi/SCR/CCP domain / Sushi/CCP/SCR domain profile. / Sushi/SCR/CCP superfamily / Metallopeptidase, catalytic domain superfamily / Neutral zinc metallopeptidases, zinc-binding region signature. / Concanavalin A-like lectin/glucanase domain superfamily
Similarity search - Domain/homology
Stanniocalcin-2 / Pappalysin-1
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.06 Å
AuthorsKobbero SD / Gajhede M / Mirza OA / Boesen T / Oxvig C
Funding support Denmark, 1 items
OrganizationGrant numberCountry
Danish Council for Independent Research Denmark
CitationJournal: Nat Commun / Year: 2022
Title: Structure of the proteolytic enzyme PAPP-A with the endogenous inhibitor stanniocalcin-2 reveals its inhibitory mechanism.
Authors: Sara Dam Kobberø / Michael Gajhede / Osman Asghar Mirza / Søren Kløverpris / Troels Rønn Kjær / Jakob Hauge Mikkelsen / Thomas Boesen / Claus Oxvig /
Abstract: The metzincin metalloproteinase PAPP-A plays a key role in the regulation of insulin-like growth factor (IGF) signaling by specific cleavage of inhibitory IGF binding proteins (IGFBPs). Using single- ...The metzincin metalloproteinase PAPP-A plays a key role in the regulation of insulin-like growth factor (IGF) signaling by specific cleavage of inhibitory IGF binding proteins (IGFBPs). Using single-particle cryo-electron microscopy (cryo-EM), we here report the structure of PAPP-A in complex with its endogenous inhibitor, stanniocalcin-2 (STC2), neither of which have been reported before. The highest resolution (3.1 Å) was obtained for the STC2 subunit and the N-terminal approximately 1000 residues of the PAPP-A subunit. The 500 kDa 2:2 PAPP-A·STC2 complex is a flexible multidomain ensemble with numerous interdomain contacts. In particular, a specific disulfide bond between the subunits of STC2 and PAPP-A prevents dissociation, and interactions between STC2 and a module located in the very C-terminal end of the PAPP-A subunit prevent binding of its main substrate, IGFBP-4. While devoid of activity towards IGFBP-4, the active site cleft of the catalytic domain is accessible in the inhibited PAPP-A·STC2 complex, as shown by its ability to hydrolyze a synthetic peptide derived from IGFBP-4. Relevant to multiple human pathologies, this unusual mechanism of proteolytic inhibition may support the development of specific pharmaceutical agents, by which IGF signaling can be indirectly modulated.
History
DepositionJun 20, 2022-
Header (metadata) releaseNov 2, 2022-
Map releaseNov 2, 2022-
UpdateNov 2, 2022-
Current statusNov 2, 2022Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15220.png

Downloads & links

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Map

FileDownload / File: emd_15220.map.gz / Format: CCP4 / Size: 40.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationPrimary, sharpened map of a partial dimer of PAPP-A in complex with a STC2 subunit
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.01 Å/pix.
x 220 pix.
= 223.08 Å		1.01 Å/pix.
x 220 pix.
= 223.08 Å		1.01 Å/pix.
x 220 pix.
= 223.08 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.014 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.05
Minimum - Maximum-0.79375446 - 1.1501384
Average (Standard dev.)0.0021747176 (±0.02685033)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions220220220
Spacing220220220
CellA=B=C: 223.08002 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Raw map of a partial dimer of PAPP-A in complex with a STC2 subunit

Fileemd_15220_additional_1.map
AnnotationRaw map of a partial dimer of PAPP-A in complex with a STC2 subunit
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A of a partial dimer of...

Fileemd_15220_half_map_1.map
AnnotationHalf map A of a partial dimer of PAPP-A in complex with a STC2 subunit
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B of a partial dimer of...

Fileemd_15220_half_map_2.map
AnnotationHalf map B of a partial dimer of PAPP-A in complex with a STC2 subunit
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Partial PAPP-A dimer in complex with subunit of its endogenous in...

EntireName: Partial PAPP-A dimer in complex with subunit of its endogenous inhibitor STC2 dimer
Components
  • Complex: Partial PAPP-A dimer in complex with subunit of its endogenous inhibitor STC2 dimer
    • Complex: PAPP-A
      • Complex: N-terminal part of PAPP-A
        • Protein or peptide: Pappalysin-1
      • Complex: C-terminal part of PAPP-A
    • Complex: Subunit of Stanniocalcin-2
      • Protein or peptide: Stanniocalcin-2
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: ZINC ION
  • Ligand: CALCIUM IONCalcium

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Supramolecule #1: Partial PAPP-A dimer in complex with subunit of its endogenous in...

SupramoleculeName: Partial PAPP-A dimer in complex with subunit of its endogenous inhibitor STC2 dimer
type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Details: Inhibited proteolytic complex generated by harvest of serum media, purifying on a nickel column followed by negative affinity purification and size-exclusion chromatography.
Source (natural)Organism: Homo sapiens (human) / Tissue: ubiquitous / Location in cell: extracellular
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: embryonic kidney cells / Recombinant plasmid: pcDNA
Molecular weightExperimental: 500 KDa

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Supramolecule #2: PAPP-A

SupramoleculeName: PAPP-A / type: complex / Chimera: Yes / ID: 2 / Parent: 1 / Macromolecule list: #1 / Details: Parts of both PAPP-A subunits
Source (natural)Organism: Homo sapiens (human) / Tissue: ubiquitous / Location in cell: extracellular
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: embryonic kidney cells / Recombinant plasmid: pcDNA

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Supramolecule #3: Subunit of Stanniocalcin-2

SupramoleculeName: Subunit of Stanniocalcin-2 / type: complex / Chimera: Yes / ID: 3 / Parent: 1 / Macromolecule list: #2 / Details: One subunit of the STC2 dimer
Source (natural)Organism: Homo sapiens (human) / Tissue: ubiquitous / Location in cell: extracellular
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: embryonic kidney cells / Recombinant plasmid: pcDNA

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Supramolecule #4: N-terminal part of PAPP-A

SupramoleculeName: N-terminal part of PAPP-A / type: complex / Chimera: Yes / ID: 4 / Parent: 2 / Macromolecule list: #1
Details: The first approximately 1000 residues of a PAPP-A subunit
Source (natural)Organism: Homo sapiens (human) / Tissue: ubiquitous / Location in cell: extracellular
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: embryonic kidney cells / Recombinant plasmid: pcDNA

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Supramolecule #5: C-terminal part of PAPP-A

SupramoleculeName: C-terminal part of PAPP-A / type: complex / Chimera: Yes / ID: 5 / Parent: 2 / Macromolecule list: #1 / Details: The C-terminal 240 residues of one PAPP-A subunit
Source (natural)Organism: Homo sapiens (human) / Tissue: ubiquitous / Location in cell: extracellular
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: embryonic kidney cells / Recombinant plasmid: pcDNA

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Macromolecule #1: Pappalysin-1

MacromoleculeName: Pappalysin-1 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: pappalysin-1
Source (natural)Organism: Homo sapiens (human) / Cell: extracellular
Molecular weightTheoretical: 171.143047 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: EARGATEEPS PPSRALYFSG RGEQLRLRAD LELPRDAFTL QVWLRAEGGQ RSPAVITGLY DKCSYISRDR GWVVGIHTIS DQDNKDPRY FFSLKTDRAR QVTTINAHRS YLPGQWVYLA ATYDGQFMKL YVNGAQVATS GEQVGGIFSP LTQKCKVLML G GSALNHNY ...String:
EARGATEEPS PPSRALYFSG RGEQLRLRAD LELPRDAFTL QVWLRAEGGQ RSPAVITGLY DKCSYISRDR GWVVGIHTIS DQDNKDPRY FFSLKTDRAR QVTTINAHRS YLPGQWVYLA ATYDGQFMKL YVNGAQVATS GEQVGGIFSP LTQKCKVLML G GSALNHNY RGYIEHFSLW KVARTQREIL SDMETHGAHT ALPQLLLQEN WDNVKHAWSP MKDGSSPKVE FSNAHGFLLD TS LEPPLCG QTLCDNTEVI ASYNQLSSFR QPKVVRYRVV NLYEDDHKNP TVTREQVDFQ HHQLAEAFKQ YNISWELDVL EVS NSSLRR RLILANCDIS KIGDENCDPE CNHTLTGHDG GDCRHLRHPA FVKKQHNGVC DMDCNYERFN FDGGECCDPE ITNV TQTCF DPDSPHRAYL DVNELKNILK LDGSTHLNIF FAKSSEEELA GVATWPWDKE ALMHLGGIVL NPSFYGMPGH THTMI HQIG HSLGLYHVFR GISEIQSCSD PCMETEPSFE TGDLCNDTNP APKHKSCGDP GPGNDTCGFH SFFNTPYNNF MSYADD DCT DSFTPNQVAR MHCYLDLVYQ GWQPSRKPAP VALAPQVLGH TTDSVTLEWF PPIDGHFFER ELGSACHLCL EGRILVQ YA SNASSPMPCS PSGHWSPREA EGHPDVEQPC KSSVRTWSPN SAVNPHTVPP ACPEPQGCYL ELEFLYPLVP ESLTIWVT F VSTDWDSSGA VNDIKLLAVS GKNISLGPQN VFCDVPLTIR LWDVGEEVYG IQIYTLDEHL EIDAAMLTST ADTPLCLQC KPLKYKVVRD PPLQMDVASI LHLNRKFVDM DLNLGSVYQY WVITISGTEE SEPSPAVTYI HGSGYCGDGI IQKDQGEQCD DMNKINGDG CSLFCRQEVS FNCIDEPSRC YFHDGDGVCE EFEQKTSIKD CGVYTPQGFL DQWASNASVS HQDQQCPGWV I IGQPAASQ VCRTKVIDLS EGISQHAWYP CTISYPYSQL AQTTFWLRAY FSQPMVAAAV IVHLVTDGTY YGDQKQETIS VQ LLDTKDQ SHDLGLHVLS CRNNPLIIPV VHDLSQPFYH SQAVRVSFSS PLVAISGVAL RSFDNFDPVT LSSCQRGETY SPA EQSCVH FACEKTDCPE LAVENASLNC SSSDRYHGAQ CTVSCRTGYV LQIRRDDELI KSQTGPSVTV TCTEGKWNKQ VACE PVDCS IPDHHQVYAA SFSCPEGTTF GSQCSFQCRH PAQLKGNNSL LTCMEDGLWS FPEALCELMC LAPPPVPNAD LQTAR CREN KHKVGSFCKY KCKPGYHVPG SSRKSKKRAF KTQCTQDGSW QEGACVPVTC DPPPPKFHGL YQCTNGFQFN SECRIK CED SDASQGLGSN VIHCRKDGTW NGSFHVCQEM QGQCSVPNEL NSNLKLQCPD GYAIGSECAT SCLDHNSESI ILPMNVT VR DIPHWLNPTR VERVVCTAGL KWYPHPALIH CVKGCEPFMG DNYCDAINNR AFCNYDGGDC CTSTVKTKKV TPFPMSCD L QGDCACRDPQ AQEHS

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Macromolecule #2: Stanniocalcin-2

MacromoleculeName: Stanniocalcin-2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human) / Cell: Extracellular
Molecular weightTheoretical: 18.819811 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
RLSLQNTAEI QHCLVNAGDV GCGVFECFEN NSCEIRGLHG ICMTFLHNAG KFDAQGKSFI KDALKCKAHA LRHRFGCISR KCPAIREMV SQLQRECYLK HDLCAAAQEN TRVIVEMIHF KDLLLHEPYV DLVNLLLTCG EEVKEAITHS VQVQCEQNWG S LCSILSF

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Macromolecule #3: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 3 / Number of copies: 8 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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Macromolecule #4: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 4 / Number of copies: 1 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #5: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 5 / Number of copies: 8 / Formula: CA
Molecular weightTheoretical: 40.078 Da

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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

Concentration0.6 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
20.0 mMHepesHepes
100.0 mMNaClSodium chlorideNaCLSodium chloride
1.0 mMCaClCaCl

Details: Hepes buffer, 20 mM Hepes pH 7.4 100 mM NaCl, 1 mM CaCl
GridModel: C-flat-2/2 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 277.15 K / Instrument: LEICA EM GP / Details: 4 s.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.6 µm
Image recording#0 - Image recording ID: 1 / #0 - Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / #0 - Number grids imaged: 1 / #0 - Number real images: 32144 / #0 - Average exposure time: 0.91 sec. / #0 - Average electron dose: 59.0 e/Å2 / #1 - Image recording ID: 2 / #1 - Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / #1 - Number grids imaged: 1 / #1 - Number real images: 10060 / #1 - Average exposure time: 0.8 sec. / #1 - Average electron dose: 58.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 6677370 / Details: Combined two datasets
CTF correctionSoftware - Name: cryoSPARC (ver. 3.3)
Initial angle assignmentType: OTHER
Final 3D classificationNumber classes: 5
Final angle assignmentType: OTHER
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.06 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3) / Number images used: 278982
Image recording ID1
FSC plot (resolution estimation)

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Atomic model buiding 1

Detailsalphafold2 model PAPP-A AF-Q13219-F1 STC2 AF-O76061-F1
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8a7d:
Partial dimer complex of PAPP-A and its inhibitor STC2

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