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- PDB-8ovb: Human Complement C3b in complex with Trypanosoma brucei ISG65. -

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Basic information

Entry
Database: PDB / ID: 8ovb
TitleHuman Complement C3b in complex with Trypanosoma brucei ISG65.
Components
  • Complement C3Complement component 3
  • Complement C3f fragment
  • ISG65 G
KeywordsIMMUNE SYSTEM / Complement system / parasite virulence / trypanosome surface protein / host-pathogen complex
Function / homology
Function and homology information


oviduct epithelium development / C5L2 anaphylatoxin chemotactic receptor binding / regulation of triglyceride biosynthetic process / positive regulation of activation of membrane attack complex / vertebrate eye-specific patterning / positive regulation of apoptotic cell clearance / complement-mediated synapse pruning / Alternative complement activation / positive regulation of lipid storage / positive regulation of G protein-coupled receptor signaling pathway ...oviduct epithelium development / C5L2 anaphylatoxin chemotactic receptor binding / regulation of triglyceride biosynthetic process / positive regulation of activation of membrane attack complex / vertebrate eye-specific patterning / positive regulation of apoptotic cell clearance / complement-mediated synapse pruning / Alternative complement activation / positive regulation of lipid storage / positive regulation of G protein-coupled receptor signaling pathway / positive regulation of phagocytosis, engulfment / complement receptor mediated signaling pathway / Activation of C3 and C5 / positive regulation of type IIa hypersensitivity / positive regulation of glucose transmembrane transport / complement-dependent cytotoxicity / complement activation, alternative pathway / complement activation / neuron remodeling / endopeptidase inhibitor activity / amyloid-beta clearance / positive regulation of vascular endothelial growth factor production / Purinergic signaling in leishmaniasis infection / complement activation, classical pathway / Peptide ligand-binding receptors / fatty acid metabolic process / Regulation of Complement cascade / response to bacterium / Post-translational protein phosphorylation / positive regulation of receptor-mediated endocytosis / positive regulation of angiogenesis / Regulation of Insulin-like Growth Factor (IGF) transport and uptake by Insulin-like Growth Factor Binding Proteins (IGFBPs) / azurophil granule lumen / Immunoregulatory interactions between a Lymphoid and a non-Lymphoid cell / G alpha (i) signalling events / secretory granule lumen / blood microparticle / immune response / inflammatory response / positive regulation of protein phosphorylation / G protein-coupled receptor signaling pathway / endoplasmic reticulum lumen / signaling receptor binding / Neutrophil degranulation / cell surface / signal transduction / protein-containing complex / extracellular space / extracellular exosome / extracellular region / membrane / plasma membrane
Similarity search - Function
Trypanosome invariant surface glycoprotein / Invariant surface glycoprotein / : / : / Complement component 3, CUB domain 2 / Complement component 3, CUB domain 1 / Complement C3-like, NTR domain / Alpha-2-macroglobulin, conserved site / Alpha-2-macroglobulin family thiolester region signature. / Complement C3/4/5, macroglobulin domain MG1 ...Trypanosome invariant surface glycoprotein / Invariant surface glycoprotein / : / : / Complement component 3, CUB domain 2 / Complement component 3, CUB domain 1 / Complement C3-like, NTR domain / Alpha-2-macroglobulin, conserved site / Alpha-2-macroglobulin family thiolester region signature. / Complement C3/4/5, macroglobulin domain MG1 / Macroglobulin domain MG1 / : / Alpha-macro-globulin thiol-ester bond-forming region / Anaphylatoxin, complement system domain / Anaphylatoxin domain signature. / Anaphylatoxin/fibulin / Anaphylatoxin, complement system / Anaphylotoxin-like domain / Anaphylatoxin domain profile. / Anaphylatoxin homologous domain / Netrin C-terminal Domain / Netrin module, non-TIMP type / UNC-6/NTR/C345C module / 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 / Netrin domain / NTR domain profile. / Tissue inhibitor of metalloproteinases-like, OB-fold / 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-like fold
Similarity search - Domain/homology
ISG65 G / Complement C3
Similarity search - Component
Biological speciesTrypanosoma brucei brucei (eukaryote)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsCook, A.D. / Higgins, M.K.
Funding support United Kingdom, 1items
OrganizationGrant numberCountry
Wellcome Trust217138/Z/19/Z United Kingdom
CitationJournal: Elife / Year: 2024
Title: Molecular mechanism of complement inhibition by the trypanosome receptor ISG65.
Authors: Alexander D Cook / Mark Carrington / Matthew K Higgins /
Abstract: African trypanosomes replicate within infected mammals where they are exposed to the complement system. This system centres around complement C3, which is present in a soluble form in serum but ...African trypanosomes replicate within infected mammals where they are exposed to the complement system. This system centres around complement C3, which is present in a soluble form in serum but becomes covalently deposited onto the surfaces of pathogens after proteolytic cleavage to C3b. Membrane-associated C3b triggers different complement-mediated effectors which promote pathogen clearance. To counter complement-mediated clearance, African trypanosomes have a cell surface receptor, ISG65, which binds to C3b and which decreases the rate of trypanosome clearance in an infection model. However, the mechanism by which ISG65 reduces C3b function has not been determined. We reveal through cryogenic electron microscopy that ISG65 has two distinct binding sites for C3b, only one of which is available in C3 and C3d. We show that ISG65 does not block the formation of C3b or the function of the C3 convertase which catalyses the surface deposition of C3b. However, we show that ISG65 forms a specific conjugate with C3b, perhaps acting as a decoy. ISG65 also occludes the binding sites for complement receptors 2 and 3, which may disrupt recruitment of immune cells, including B cells, phagocytes, and granulocytes. This suggests that ISG65 protects trypanosomes by combining multiple approaches to dampen the complement cascade.
History
DepositionApr 25, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 14, 2024Provider: repository / Type: Initial release
Revision 1.1May 15, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Complement C3f fragment
B: Complement C3
C: ISG65 G


Theoretical massNumber of molelcules
Total (without water)207,8253
Polymers207,8253
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: surface plasmon resonance
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Complement C3f fragment


Mass: 71154.047 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Tissue: Blood / References: UniProt: P01024
#2: Protein Complement C3 / Complement component 3 / C3 and PZP-like alpha-2-macroglobulin domain-containing protein 1


Mass: 104073.164 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Tissue: Blood / References: UniProt: P01024
#3: Protein ISG65 G


Mass: 32597.322 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Trypanosoma brucei brucei (eukaryote) / Gene: ISG65 G / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: A0A8J9S0Z8

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSourceDetails
1Complex of Human Complement C3b and T. brucei ISG65COMPLEXall0MULTIPLE SOURCES
2Human Complement C3bCOMPLEX#1-#21NATURALC3 purified from human serum, C3b generated from C3 by limited trypsin proteolysis.
3Trypanosoma brucei ISG65COMPLEX#31RECOMBINANT
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
110.212 MDaNO
210.176 MDaNO
310.0442 MDaNO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-IDTissue
22Homo sapiens (human)9606Blood
32Homo sapiens (human)9606Blood
43Trypanosoma brucei brucei (eukaryote)5702
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPES1
2150 mMSaltNaClSodium chloride1
SpecimenConc.: 2.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 278.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 3.03 sec. / Electron dose: 49.9 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 14339
EM imaging opticsEnergyfilter name: GIF Quantum LS / Energyfilter slit width: 20 eV

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Processing

EM software
IDNameVersionCategory
1SIMPLE3particle selection
2cryoSPARC3particle selection
3RELION3.1particle selection
4EPUimage acquisition
6SIMPLE3CTF correction
9UCSF ChimeraXmodel fitting
11SIMPLE3initial Euler assignment
12cryoSPARC3initial Euler assignment
13cryoSPARCfinal Euler assignment
14cryoSPARC3classification
15cryoSPARC33D reconstruction
16ISOLDEmodel refinement
17Cootmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 3824878
3D reconstructionResolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 481606 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: REAL
Atomic model building
IDPDB-ID 3D fitting-IDSource nameTypeAccession codeInitial refinement model-ID
11AlphaFoldin silico model
25FO7

5fo7

1PDBexperimental model5FO72

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