+Open data
-Basic information
Entry | Database: PDB / ID: 7kwo | ||||||
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Title | rFVIIIFc-VWF-XTEN (BIVV001) | ||||||
Components |
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Keywords | BLOOD CLOTTING / Complex / Hemophilia / FVIII / VWF | ||||||
Function / homology | Function and homology information Defective F8 accelerates dissociation of the A2 domain / Defective F8 binding to the cell membrane / Defective F8 secretion / Defective F8 sulfation at Y1699 / Gamma carboxylation, hypusinylation, hydroxylation, and arylsulfatase activation / Defective VWF binding to collagen type I / Enhanced cleavage of VWF variant by ADAMTS13 / Defective VWF cleavage by ADAMTS13 variant / Defective F8 binding to von Willebrand factor / hemostasis ...Defective F8 accelerates dissociation of the A2 domain / Defective F8 binding to the cell membrane / Defective F8 secretion / Defective F8 sulfation at Y1699 / Gamma carboxylation, hypusinylation, hydroxylation, and arylsulfatase activation / Defective VWF binding to collagen type I / Enhanced cleavage of VWF variant by ADAMTS13 / Defective VWF cleavage by ADAMTS13 variant / Defective F8 binding to von Willebrand factor / hemostasis / Enhanced binding of GP1BA variant to VWF multimer:collagen / Defective binding of VWF variant to GPIb:IX:V / Weibel-Palade body / blood coagulation, intrinsic pathway / platelet alpha granule / Platelet Adhesion to exposed collagen / Cargo concentration in the ER / Defective factor IX causes thrombophilia / Defective cofactor function of FVIIIa variant / Defective F9 variant does not activate FX / positive regulation of intracellular signal transduction / COPII-mediated vesicle transport / GP1b-IX-V activation signalling / COPII-coated ER to Golgi transport vesicle / p130Cas linkage to MAPK signaling for integrins / cell-substrate adhesion / Defective F8 cleavage by thrombin / Platelet Aggregation (Plug Formation) / immunoglobulin binding / GRB2:SOS provides linkage to MAPK signaling for Integrins / Integrin cell surface interactions / Common Pathway of Fibrin Clot Formation / collagen binding / Intrinsic Pathway of Fibrin Clot Formation / endoplasmic reticulum-Golgi intermediate compartment membrane / Integrin signaling / extracellular matrix / platelet alpha granule lumen / acute-phase response / Signaling by high-kinase activity BRAF mutants / MAP2K and MAPK activation / platelet activation / response to wounding / Golgi lumen / Signaling by RAF1 mutants / Signaling by moderate kinase activity BRAF mutants / Paradoxical activation of RAF signaling by kinase inactive BRAF / Signaling downstream of RAS mutants / Signaling by BRAF and RAF1 fusions / blood coagulation / integrin binding / Platelet degranulation / protein-folding chaperone binding / protease binding / collagen-containing extracellular matrix / oxidoreductase activity / cell adhesion / copper ion binding / endoplasmic reticulum lumen / endoplasmic reticulum / extracellular space / extracellular exosome / extracellular region / identical protein binding / plasma membrane Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å | ||||||
Authors | Fuller, J.R. / Batchelor, J.D. | ||||||
Citation | Journal: Blood / Year: 2021 Title: Molecular determinants of the factor VIII/von Willebrand factor complex revealed by BIVV001 cryo-electron microscopy. Authors: James R Fuller / Kevin E Knockenhauer / Nina C Leksa / Robert T Peters / Joseph D Batchelor Abstract: Interaction of factor VIII (FVIII) with von Willebrand factor (VWF) is mediated by the VWF D'D3 domains and thrombin-mediated release is essential for hemostasis after vascular injury. VWF-D'D3 ...Interaction of factor VIII (FVIII) with von Willebrand factor (VWF) is mediated by the VWF D'D3 domains and thrombin-mediated release is essential for hemostasis after vascular injury. VWF-D'D3 mutations resulting in loss of FVIII binding are the underlying cause of von Willebrand disease (VWD) type 2N. Furthermore, the FVIII-VWF interaction has significant implications for the development of therapeutics for bleeding disorders, particularly hemophilia A, in which endogenous VWF clearance imposes a half-life ceiling on replacement FVIII therapy. To understand the structural basis of FVIII engagement by VWF, we solved the structure of BIVV001 by cryo-electron microscopy to 2.9 Å resolution. BIVV001 is a bioengineered clinical-stage FVIII molecule for the treatment of hemophilia A. In BIVV001, VWF-D'D3 is covalently linked to an Fc domain of a B domain-deleted recombinant FVIII (rFVIII) Fc fusion protein, resulting in a stabilized rFVIII/VWF-D'D3 complex. Our rFVIII/VWF structure resolves BIVV001 architecture and provides a detailed spatial understanding of previous biochemical and clinical observations related to FVIII-VWF engagement. Notably, the FVIII acidic a3 peptide region (FVIII-a3), established as a critical determinant of FVIII/VWF complex formation, inserts into a basic groove formed at the VWF-D'/rFVIII interface. Our structure shows direct interaction of sulfated Y1680 in FVIII-a3 and VWF-R816 that, when mutated, leads to severe hemophilia A or VWD type 2N, respectively. These results provide insight on this key coagulation complex, explain the structural basis of many hemophilia A and VWD type 2N mutations, and inform studies to further elucidate how VWF dissociates rapidly from FVIII upon activation. #1: Journal: Comput. Cryst. Newsl. / Year: 2013 Title: New tool: phenix.real_space_refine Authors: Afonine, P.V. / Headd, J.J. / Terwilliger, T.C. / Adams, P.D. #2: Journal: Acta Crystallogr D Struct Biol / Year: 2019 Title: Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix. Authors: Dorothee Liebschner / Pavel V Afonine / Matthew L Baker / Gábor Bunkóczi / Vincent B Chen / Tristan I Croll / Bradley Hintze / Li Wei Hung / Swati Jain / Airlie J McCoy / Nigel W Moriarty ...Authors: Dorothee Liebschner / Pavel V Afonine / Matthew L Baker / Gábor Bunkóczi / Vincent B Chen / Tristan I Croll / Bradley Hintze / Li Wei Hung / Swati Jain / Airlie J McCoy / Nigel W Moriarty / Robert D Oeffner / Billy K Poon / Michael G Prisant / Randy J Read / Jane S Richardson / David C Richardson / Massimo D Sammito / Oleg V Sobolev / Duncan H Stockwell / Thomas C Terwilliger / Alexandre G Urzhumtsev / Lizbeth L Videau / Christopher J Williams / Paul D Adams / Abstract: Diffraction (X-ray, neutron and electron) and electron cryo-microscopy are powerful methods to determine three-dimensional macromolecular structures, which are required to understand biological ...Diffraction (X-ray, neutron and electron) and electron cryo-microscopy are powerful methods to determine three-dimensional macromolecular structures, which are required to understand biological processes and to develop new therapeutics against diseases. The overall structure-solution workflow is similar for these techniques, but nuances exist because the properties of the reduced experimental data are different. Software tools for structure determination should therefore be tailored for each method. Phenix is a comprehensive software package for macromolecular structure determination that handles data from any of these techniques. Tasks performed with Phenix include data-quality assessment, map improvement, model building, the validation/rebuilding/refinement cycle and deposition. Each tool caters to the type of experimental data. The design of Phenix emphasizes the automation of procedures, where possible, to minimize repetitive and time-consuming manual tasks, while default parameters are chosen to encourage best practice. A graphical user interface provides access to many command-line features of Phenix and streamlines the transition between programs, project tracking and re-running of previous tasks. #3: Journal: Acta Crystallogr D Struct Biol / Year: 2018 Title: ISOLDE: a physically realistic environment for model building into low-resolution electron-density maps. Authors: Tristan Ian Croll / Abstract: This paper introduces ISOLDE, a new software package designed to provide an intuitive environment for high-fidelity interactive remodelling/refinement of macromolecular models into electron-density ...This paper introduces ISOLDE, a new software package designed to provide an intuitive environment for high-fidelity interactive remodelling/refinement of macromolecular models into electron-density maps. ISOLDE combines interactive molecular-dynamics flexible fitting with modern molecular-graphics visualization and established structural biology libraries to provide an immersive interface wherein the model constantly acts to maintain physically realistic conformations as the user interacts with it by directly tugging atoms with a mouse or haptic interface or applying/removing restraints. In addition, common validation tasks are accelerated and visualized in real time. Using the recently described 3.8 Å resolution cryo-EM structure of the eukaryotic minichromosome maintenance (MCM) helicase complex as a case study, it is demonstrated how ISOLDE can be used alongside other modern refinement tools to avoid common pitfalls of low-resolution modelling and improve the quality of the final model. A detailed analysis of changes between the initial and final model provides a somewhat sobering insight into the dangers of relying on a small number of validation metrics to judge the quality of a low-resolution model. #4: Journal: Elife / Year: 2016 Title: Automated structure refinement of macromolecular assemblies from cryo-EM maps using Rosetta. Authors: Ray Yu-Ruei Wang / Yifan Song / Benjamin A Barad / Yifan Cheng / James S Fraser / Frank DiMaio / Abstract: Cryo-EM has revealed the structures of many challenging yet exciting macromolecular assemblies at near-atomic resolution (3-4.5Å), providing biological phenomena with molecular descriptions. ...Cryo-EM has revealed the structures of many challenging yet exciting macromolecular assemblies at near-atomic resolution (3-4.5Å), providing biological phenomena with molecular descriptions. However, at these resolutions, accurately positioning individual atoms remains challenging and error-prone. Manually refining thousands of amino acids - typical in a macromolecular assembly - is tedious and time-consuming. We present an automated method that can improve the atomic details in models that are manually built in near-atomic-resolution cryo-EM maps. Applying the method to three systems recently solved by cryo-EM, we are able to improve model geometry while maintaining the fit-to-density. Backbone placement errors are automatically detected and corrected, and the refinement shows a large radius of convergence. The results demonstrate that the method is amenable to structures with symmetry, of very large size, and containing RNA as well as covalently bound ligands. The method should streamline the cryo-EM structure determination process, providing accurate and unbiased atomic structure interpretation of such maps. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7kwo.cif.gz | 361.5 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7kwo.ent.gz | 255 KB | Display | PDB format |
PDBx/mmJSON format | 7kwo.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7kwo_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 7kwo_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 7kwo_validation.xml.gz | 63.6 KB | Display | |
Data in CIF | 7kwo_validation.cif.gz | 95 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/kw/7kwo ftp://data.pdbj.org/pub/pdb/validation_reports/kw/7kwo | HTTPS FTP |
-Related structure data
Related structure data | 23057MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
-Protein , 2 types, 2 molecules AV
#1: Protein | Mass: 218874.969 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Production host: Homo sapiens (human) / References: UniProt: P00451*PLUS |
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#2: Protein | Mass: 179048.953 Da / Num. of mol.: 1 / Mutation: C1099A, C1142A Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Production host: Homo sapiens (human) / References: UniProt: P04275*PLUS |
-Sugars , 2 types, 4 molecules
#3: Polysaccharide | beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source |
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#4: Sugar |
-Non-polymers , 3 types, 4 molecules
#5: Chemical | #6: Chemical | ChemComp-ZN / | #7: Chemical | ChemComp-CU / | |
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-Details
Compound details | Coagulation factor FVIII-Fc-XTEN is fusion protein of human coagulation factor VIII (Uniprot: ...Coagulation factor FVIII-Fc-XTEN is fusion protein of human coagulation factor VIII (Uniprot: P00451), XTEN polypeptide, and Immunoglobulin heavy constant gamma 1 (UniProt: P01857). von Willebrand factor-XTEN-Fc is fusion protein of human von Willebrand factor (VWF) (Uniprot: P04275), XTEN polypeptide, and Immunoglobulin heavy constant gamma 1 (UniProt: P01857). |
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Has ligand of interest | Y |
Has protein modification | Y |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: rFVIIIFc-VWF-XTEN (BIVV001) / Type: COMPLEX Details: An engineered therapeutic complex between coagulation factor VIII and von Willebrand factor D'D3 Entity ID: #1-#2 / Source: RECOMBINANT | |||||||||||||||||||||||||||||||||||||||||||||
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Molecular weight | Value: 0.31164974 MDa / Experimental value: NO | |||||||||||||||||||||||||||||||||||||||||||||
Source (natural) | Organism: Homo sapiens (human) | |||||||||||||||||||||||||||||||||||||||||||||
Source (recombinant) | Organism: Homo sapiens (human) | |||||||||||||||||||||||||||||||||||||||||||||
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Specimen | Experiment-ID: 1 / Conc.: 0.75 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Prepared by gel filtration chromatography immediately prior to vitrification.
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Specimen support |
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Vitrification |
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-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 81000 X / Nominal defocus max: 2600 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
Image scans | Width: 11520 / Height: 8184 |
-Processing
Software |
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EM software |
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Image processing | Details: Micrograph movies were motion-corrected and dose-weighted using Relion 3.0 | ||||||||||||||||||||||||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 116200 / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL | ||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | 3D fitting-ID: 1 / Source name: PDB / Type: experimental model
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Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 32.14 Å2 | ||||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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