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- EMDB-71094: Nanodisc-embedded human TF/FVIIa/XK1 -

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Basic information

Entry
Database: EMDB / ID: EMD-71094
TitleNanodisc-embedded human TF/FVIIa/XK1
Map dataNanodisc-embedded human TF/FVIIa/XK1
Sample
  • Complex: Complex of factor VIIa and XK1, bound to tissue factor embedded in a nanodisc membrane bilayer.
    • Complex: XK1 chimeric protein
      • Protein or peptide: Human factor X (FX) light chain
      • Protein or peptide: Human TFPI Kunitz domain 1 (K1)
    • Complex: Human factor VIIa (FVIIa)
      • Protein or peptide: Human factor VIIa - light chain
      • Protein or peptide: Human factor VIIa (FVIIa) protease domain
    • Complex: Human MBP-tagged tissue factor
      • Protein or peptide: Human tissue factor (TF)
Keywordscomplex / nanodisc / blood clotting
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.0 Å
AuthorsPhotenhauer AL / Sedzro JC / Ohi MD / Morrissey JH
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)R35 HL135823 United States
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)R35 HL171334 United States
National Institutes of Health/Office of the DirectorS10OD030275 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R24 GM145965 United States
American Heart AssociationN028347 United States
CitationJournal: Blood / Year: 2025
Title: Cryo-EM structure of the tissue factor/factor VIIa complex with a factor X mimetic reveals a novel allosteric mechanism.
Authors: Josepha C Sedzro / Amanda L Photenhauer / Fabienne Birkle / Katarina Meze / Alex Mortenson / Cade Duckworth / Po-Chao Wen / Sarah Kearns / Michael A Cianfrocco / Emad Tajkhorshid / Melanie D ...Authors: Josepha C Sedzro / Amanda L Photenhauer / Fabienne Birkle / Katarina Meze / Alex Mortenson / Cade Duckworth / Po-Chao Wen / Sarah Kearns / Michael A Cianfrocco / Emad Tajkhorshid / Melanie D Ohi / James H Morrissey /
Abstract: Blood clotting is triggered in hemostasis and thrombosis when the membrane-bound tissue factor (TF)/factor VIIa (FVIIa) complex activates factor X (FX). There are no structures of TF/FVIIa on ...Blood clotting is triggered in hemostasis and thrombosis when the membrane-bound tissue factor (TF)/factor VIIa (FVIIa) complex activates factor X (FX). There are no structures of TF/FVIIa on membranes, with or without FX. Using cryo-EM to address this gap, we assembled TF/FVIIa complexes on nanoscale membrane bilayers (nanodiscs), bound to XK1 and an antibody fragment. XK1 is a FX mimetic whose protease domain is replaced by the first Kunitz-type (K1) domain of tissue factor pathway inhibitor, while 10H10 is a non-inhibitory, anti-TF antibody. We determined a cryo-EM structure of a TF/FVIIa/XK1/10H10/nanodisc complex with a resolution of 3.7 Å, allowing us to model all the protein backbones. TF/FVIIa extends perpendicularly from the membrane, interacting with a "handle shaped" XK1 at two locations: the K1 domain docks into FVIIa's active site, while the γ-carboxyglutamate-rich (GLA) domain binds to TF's substrate-binding exosite. The FX and FVIIa GLA domains also contact each other and the membrane surface. Except for a minor contact between the first epidermal growth factor (EGF)-like domain of XK1 and TF, the rest of the FX light chain does not interact with TF/FVIIa. The structure reveals a previously unrecognized, membrane-dependent allosteric activation mechanism between FVIIa and TF where a serine-rich loop in TF that partially obscures the TF exosite must undergo a shift to allow access of the FX GLA domain to its final binding location on the membrane-bound TF/FVIIa complex. This mechanism also provides a novel explanation for the otherwise puzzling phenomenon of TF encryption/decryption on cell surfaces.
History
DepositionJun 9, 2025-
Header (metadata) releaseAug 27, 2025-
Map releaseAug 27, 2025-
UpdateAug 27, 2025-
Current statusAug 27, 2025Processing site: RCSB / Status: Released

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Structure visualization

Downloads & links

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Map

FileDownload / File: emd_71094.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNanodisc-embedded human TF/FVIIa/XK1
Voxel sizeX=Y=Z: 0.834 Å
Density
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-1.5739081 - 1.9860791
Average (Standard dev.)-0.000733861 (±0.056037568)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 333.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Complex of factor VIIa and XK1, bound to tissue factor embedded i...

EntireName: Complex of factor VIIa and XK1, bound to tissue factor embedded in a nanodisc membrane bilayer.
Components
  • Complex: Complex of factor VIIa and XK1, bound to tissue factor embedded in a nanodisc membrane bilayer.
    • Complex: XK1 chimeric protein
      • Protein or peptide: Human factor X (FX) light chain
      • Protein or peptide: Human TFPI Kunitz domain 1 (K1)
    • Complex: Human factor VIIa (FVIIa)
      • Protein or peptide: Human factor VIIa - light chain
      • Protein or peptide: Human factor VIIa (FVIIa) protease domain
    • Complex: Human MBP-tagged tissue factor
      • Protein or peptide: Human tissue factor (TF)

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Supramolecule #1: Complex of factor VIIa and XK1, bound to tissue factor embedded i...

SupramoleculeName: Complex of factor VIIa and XK1, bound to tissue factor embedded in a nanodisc membrane bilayer.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all / Details: Map with no 10H10 Fab

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Supramolecule #2: XK1 chimeric protein

SupramoleculeName: XK1 chimeric protein / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #3, #5
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #3: Human factor VIIa (FVIIa)

SupramoleculeName: Human factor VIIa (FVIIa) / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #1-#2
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #4: Human MBP-tagged tissue factor

SupramoleculeName: Human MBP-tagged tissue factor / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #4
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Human factor VIIa - light chain

MacromoleculeName: Human factor VIIa - light chain / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
ANAFL(CGU)(CGU)LRP GSL(CGU)R(CGU)CK(CGU)(CGU) QCSF(CGU)(CGU)AR(CGU)I FKDA(CGU)RTKLF

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Macromolecule #2: Human factor VIIa (FVIIa) protease domain

MacromoleculeName: Human factor VIIa (FVIIa) protease domain / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
IVGGKVCPKG ECPWQVLLLV NGAQLCGGTL INTIWVVSAA HCFDKIKNWR NLIAVLGEHD LSEHDGDEQS RRVAQVIIPS

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Macromolecule #3: Human factor X (FX) light chain

MacromoleculeName: Human factor X (FX) light chain / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
ANSFL(CGU)(CGU)MKK GHL(CGU)R(CGU)CM(CGU)(CGU) TCSY(CGU)(CGU)AR(CGU)V F(CGU)DSDKTN

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Macromolecule #4: Human tissue factor (TF)

MacromoleculeName: Human tissue factor (TF) / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: TVAAYNLTWK STNFKTILEW EPKPVNQVYT VQISTKSGDW KSKCFYTTDT ECDLTDEIVK DVKQTYLARV FSYPAGNVES TGSAGEPLYE NSPEFTPYLE TNLGQPTIQS FEQVGTKVNV TVEDERTLVR RNNTFLSLRD VFGKDLIYTL YYWKSSSSGK KTAKTNTNEF ...String:
TVAAYNLTWK STNFKTILEW EPKPVNQVYT VQISTKSGDW KSKCFYTTDT ECDLTDEIVK DVKQTYLARV FSYPAGNVES TGSAGEPLYE NSPEFTPYLE TNLGQPTIQS FEQVGTKVNV TVEDERTLVR RNNTFLSLRD VFGKDLIYTL YYWKSSSSGK KTAKTNTNEF LIDVDKGENY CFSVQAVIPS RTVNRKSTDS PVECMGQEKG EFREIFYIIG AVVFVVIILV IILAISLHKG SGSGMKIEEG KLVIWINGDK GYNGLAEVGK KFEKDTGIKV TVEHPDKLEE KFPQVAATGD GPDIIFWAHD RFGGYAQSGL LAEITPDKAF QDKLYPFTWD AVRYNGKLIA YPIAVEALSL IYNKDLLPNP PKTWEEIPAL DKELKAKGKS ALMFNLQEPY FTWPLIAADG GYAFKYENGK YDIKDVGVDN AGAKAGLTFL VDLIKNKHMN ADTDYSIAEA AFNKGETAMT INGPWAWSNI DTSKVNYGVT VLPTFKGQPS KPFVGVLSAG INAASPNKEL AKEFLENYLL TDEGLEAVNK DKPLGAVALK SYEEELAKDP RIAATMENAQ KGEIMPNIPQ MSAFWYAVRT AVINAASGRQ TVDEALKDAQ T

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Macromolecule #5: Human TFPI Kunitz domain 1 (K1)

MacromoleculeName: Human TFPI Kunitz domain 1 (K1) / type: protein_or_peptide / ID: 5 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
TDTELPPLKL MHSFCAFKAD DGPCKAIMKR FFFNIFTRQC EEFIYGGCEG NQNRFESLEE CKKMCTRD

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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
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Slit width: 20 eV
SoftwareName: SerialEM
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 20088 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 105000
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1161867
CTF correctionType: NONE
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 6.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.5.1) / Software - details: Non-uniform refinement / Number images used: 96859
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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Atomic model buiding 1

Initial modelPDB ID:

9p0x


Chain - Source name: PDB / Chain - Initial model type: experimental model / Details: Model generated from associated entry

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