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- EMDB-48465: Cryo-EM structure of factor Va bound to activated protein C -

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Basic information

Entry
Database: EMDB / ID: EMD-48465
TitleCryo-EM structure of factor Va bound to activated protein C
Map dataA1 domain local refined map
Sample
  • Complex: Complex of coagulation Factor Va and Activated Protein C
    • Complex: Coagulation Factor Va
    • Complex: Activated Protein C
KeywordsCoagulation / Activated Factor V / Activated Protein C / BLOOD CLOTTING
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.0 Å
AuthorsMohammed BM / Basore K / Di Cera E
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)HL049413, HL139554 and HL147821 United States
Childrens Discovery Institute of Washington University and St. Louis Childrens HospitalCDI-CORE-2015-505 and CDI-CORE-2019-813 United States
The Foundation for Barnes-Jewish Hospital3770 United States
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)DK020579 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA091842 United States
CitationJournal: Blood / Year: 2025
Title: Cryo-EM structure of coagulation factor Va bound to activated protein C.
Authors: Bassem M Mohammed / Katherine Basore / Enrico Di Cera /
Abstract: Coagulation factor Va (fVa) is the cofactor component of the prothrombinase complex required for rapid generation of thrombin from prothrombin in the penultimate step of the coagulation cascade. In ...Coagulation factor Va (fVa) is the cofactor component of the prothrombinase complex required for rapid generation of thrombin from prothrombin in the penultimate step of the coagulation cascade. In addition, fVa is a target for proteolytic inactivation by activated protein C (APC). Like other protein-protein interactions in the coagulation cascade, the fVa-APC interaction has long posed a challenge to structural biology and its molecular underpinnings remain unknown. A recent cryogenic electron microscopy (cryo-EM) structure of fVa has revealed the arrangement of its A1-A2-A3-C1-C2 domains and the environment of the sites of APC cleavage at R306 and R506. Here we report the cryo-EM structure of the fVa-APC complex at 3.15 Å resolution where the protease domain of APC engages R506 in the A2 domain of fVa mainly through electrostatic interactions between positively charged residues in the 30- and 70- loops of APC and an electronegative surface of fVa. The auxiliary Gla and EGF domains of APC are highly dynamic and point to solvent, without making contacts with fVa. Binding of APC displaces a large portion of the A2 domain of fVa and projects the 654VKCIPDDDEDSYEIFEP670 segment as a "latch", or exosite ligand, over the 70-loop of the enzyme. The latch induces a large conformational change of the autolysis loop of APC which in turn promotes docking of R506 into the primary specificity pocket. The cryo-EM structure of the fVa-APC complex validates the bulk of existing biochemical data and offers molecular context for a key regulatory interaction of the coagulation cascade.
History
DepositionDec 26, 2024-
Header (metadata) releaseMay 21, 2025-
Map releaseMay 21, 2025-
UpdateMay 21, 2025-
Current statusMay 21, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_48465.png

Downloads & links

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Map

FileDownload / File: emd_48465.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationA1 domain local refined map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.87 Å/pix.
x 420 pix.
= 365.442 Å		0.87 Å/pix.
x 420 pix.
= 365.442 Å		0.87 Å/pix.
x 420 pix.
= 365.442 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.8701 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.234
Minimum - Maximum-1.312464 - 2.2832675
Average (Standard dev.)-0.00012879644 (±0.021463642)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions420420420
Spacing420420420
CellA=B=C: 365.44202 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_48465_msk_1.map
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Additional map: Mask for DeepEmhancer

Fileemd_48465_additional_1.map
AnnotationMask for DeepEmhancer
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Additional map: DeepEmhancer sharpened map

Fileemd_48465_additional_2.map
AnnotationDeepEmhancer sharpened map
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AxesZYX

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Half map: #1

Fileemd_48465_half_map_1.map
Projections & Slices
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Half map: #2

Fileemd_48465_half_map_2.map
Projections & Slices
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Sample components

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Entire : Complex of coagulation Factor Va and Activated Protein C

EntireName: Complex of coagulation Factor Va and Activated Protein C
Components
  • Complex: Complex of coagulation Factor Va and Activated Protein C
    • Complex: Coagulation Factor Va
    • Complex: Activated Protein C

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Supramolecule #1: Complex of coagulation Factor Va and Activated Protein C

SupramoleculeName: Complex of coagulation Factor Va and Activated Protein C
type: complex / ID: 1 / Parent: 0
Details: Coagulation activated Factor V (FVa) [from plasma] complex with Activated Protein C (APC) [made recombinantly with Ser360Ala mutation].

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Supramolecule #2: Coagulation Factor Va

SupramoleculeName: Coagulation Factor Va / type: complex / ID: 2 / Parent: 1
Source (natural)Organism: Homo sapiens (human) / Tissue: Blood

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Supramolecule #3: Activated Protein C

SupramoleculeName: Activated Protein C / type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Homo sapiens (human)

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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 #1

Preparation ID1
Concentration0.1 mg/mL
BufferpH: 7.4 / Details: 20 mM HEPES, 150 mM NaCl, 5 mM CaCl2
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Film type ID: 1 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 40 sec. / Pretreatment - Atmosphere: AIR / Details: Grids were coated with polylysine 0.1 mg/mL
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
Details0.1 mg/mL

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Sample preparation #2

Preparation ID2
Concentration0.1 mg/mL
BufferpH: 7.4 / Details: 20 mM HEPES, 150 mM NaCl, 5 mM CaCl2
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Film type ID: 1 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 40 sec. / Pretreatment - Atmosphere: AIR / Details: Grids were coated with polylysine 0.1 mg/mL
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Sample preparation #3

Preparation ID3
Concentration0.1 mg/mL
BufferpH: 7.4 / Details: 20 mM HEPES, 150 mM NaCl, 5 mM CaCl2
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Film type ID: 1 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 40 sec. / Pretreatment - Atmosphere: AIR / Details: Grids were coated with polylysine 0.1 mg/mL
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Image recording#0 - Image recording ID: 1 / #0 - Film or detector model: FEI FALCON IV (4k x 4k) / #0 - Detector mode: COUNTING / #0 - Digitization - Dimensions - Width: 4096 pixel / #0 - Digitization - Dimensions - Height: 4096 pixel / #0 - Number grids imaged: 1 / #0 - Number real images: 3193 / #0 - Average electron dose: 51.86 e/Å2 / #0 - Details: 30 degrees tilt / #1 - Image recording ID: 2 / #1 - Film or detector model: FEI FALCON IV (4k x 4k) / #1 - Digitization - Dimensions - Width: 4096 pixel / #1 - Digitization - Dimensions - Height: 4096 pixel / #1 - Number grids imaged: 1 / #1 - Number real images: 600 / #1 - Average electron dose: 46.6 e/Å2 / #2 - Image recording ID: 3 / #2 - Film or detector model: FEI FALCON IV (4k x 4k) / #2 - Digitization - Dimensions - Width: 4096 pixel / #2 - Digitization - Dimensions - Height: 4096 pixel / #2 - Number grids imaged: 1 / #2 - Number real images: 2655 / #2 - Average electron dose: 46.89 e/Å2 / #2 - Details: 30 degrees tilt / #3 - Image recording ID: 4 / #3 - Film or detector model: FEI FALCON IV (4k x 4k) / #3 - Digitization - Dimensions - Width: 4096 pixel / #3 - Digitization - Dimensions - Height: 4096 pixel / #3 - Number grids imaged: 1 / #3 - Number real images: 2379 / #3 - Average electron dose: 47.19 e/Å2 / #4 - Image recording ID: 5 / #4 - Film or detector model: FEI FALCON IV (4k x 4k) / #4 - Digitization - Dimensions - Width: 4096 pixel / #4 - Digitization - Dimensions - Height: 4096 pixel / #4 - Number grids imaged: 1 / #4 - Number real images: 420 / #4 - Average electron dose: 46.89 e/Å2 / #5 - Image recording ID: 6 / #5 - Film or detector model: FEI FALCON IV (4k x 4k) / #5 - Digitization - Dimensions - Width: 4096 pixel / #5 - Digitization - Dimensions - Height: 4096 pixel / #5 - Number grids imaged: 1 / #5 - Number real images: 3210 / #5 - Average electron dose: 52.8 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.01 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 75000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Image recording ID1
CTF correctionSoftware - Name: cryoSPARC (ver. 4.6.2) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.6.2) / Number images used: 384239
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
FSC plot (resolution estimation)

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