+Open data
-Basic information
Entry | Database: PDB / ID: 9b4h | ||||||||||||||||||||||||
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Title | Chlamydomonas reinhardtii mastigoneme filament | ||||||||||||||||||||||||
Components |
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Keywords | STRUCTURAL PROTEIN / Mastigoneme / Glycosylated hydroxyproline | ||||||||||||||||||||||||
Function / homology | Function and homology information | ||||||||||||||||||||||||
Biological species | Chlamydomonas reinhardtii (plant) | ||||||||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å | ||||||||||||||||||||||||
Authors | Dai, J. / Ma, M. / Zhang, R. / Brown, A. | ||||||||||||||||||||||||
Funding support | United States, 7items
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Citation | Journal: Cell / Year: 2024 Title: Mastigoneme structure reveals insights into the O-linked glycosylation code of native hydroxyproline-rich helices. Authors: Jin Dai / Meisheng Ma / Qingwei Niu / Robyn J Eisert / Xiangli Wang / Poulomi Das / Karl F Lechtreck / Susan K Dutcher / Rui Zhang / Alan Brown / Abstract: Hydroxyproline-rich glycoproteins (HRGPs) are a ubiquitous class of protein in the extracellular matrices and cell walls of plants and algae, yet little is known of their native structures or ...Hydroxyproline-rich glycoproteins (HRGPs) are a ubiquitous class of protein in the extracellular matrices and cell walls of plants and algae, yet little is known of their native structures or interactions. Here, we used electron cryomicroscopy (cryo-EM) to determine the structure of the hydroxyproline-rich mastigoneme, an extracellular filament isolated from the cilia of the alga Chlamydomonas reinhardtii. The structure demonstrates that mastigonemes are formed from two HRGPs (a filament of MST1 wrapped around a single copy of MST3) that both have hyperglycosylated poly(hydroxyproline) helices. Within the helices, O-linked glycosylation of the hydroxyproline residues and O-galactosylation of interspersed serine residues create a carbohydrate casing. Analysis of the associated glycans reveals how the pattern of hydroxyproline repetition determines the type and extent of glycosylation. MST3 possesses a PKD2-like transmembrane domain that forms a heteromeric polycystin-like cation channel with PKD2 and SIP, explaining how mastigonemes are tethered to ciliary membranes. | ||||||||||||||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 9b4h.cif.gz | 970.1 KB | Display | PDBx/mmCIF format |
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PDB format | pdb9b4h.ent.gz | Display | PDB format | |
PDBx/mmJSON format | 9b4h.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 9b4h_validation.pdf.gz | 6 MB | Display | wwPDB validaton report |
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Full document | 9b4h_full_validation.pdf.gz | 6.2 MB | Display | |
Data in XML | 9b4h_validation.xml.gz | 111.5 KB | Display | |
Data in CIF | 9b4h_validation.cif.gz | 173.8 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/b4/9b4h ftp://data.pdbj.org/pub/pdb/validation_reports/b4/9b4h | HTTPS FTP |
-Related structure data
Related structure data | 43892MC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Protein , 2 types, 3 molecules ABX
#1: Protein | Mass: 206316.203 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / Strain: CC-4402 / References: UniProt: A8J9H7 #2: Protein | | Mass: 868021.500 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / Strain: CC-4402 / References: UniProt: A0A2K3DRP3 |
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-Sugars , 8 types, 245 molecules
#3: Polysaccharide | 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source #4: Polysaccharide | beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose Source method: isolated from a genetically manipulated source #5: Polysaccharide | beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose-(5-5)-beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose Type: oligosaccharide / Mass: 546.474 Da / Num. of mol.: 10 Source method: isolated from a genetically manipulated source #6: Polysaccharide | beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose-(5-5)-beta-L-arabinofuranose-(1-2)-beta-L- ...beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose-(5-5)-beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose-(5-5)-beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose-(5-5)-beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose | Type: oligosaccharide / Mass: 1074.934 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source #7: Polysaccharide | beta-L-arabinofuranose-(1-4)-beta-L-ribulofuranose | Type: oligosaccharide / Mass: 282.245 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source #8: Polysaccharide | beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose-(5-5)-beta-L-arabinofuranose-(1-2)-beta-L- ...beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose-(5-5)-beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose-(5-5)-beta-L-arabinofuranose-(1-2)-beta-L-arabinofuranose | Type: oligosaccharide / Mass: 810.704 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source #9: Sugar | ChemComp-A1AIO / Type: L-saccharide / Mass: 180.156 Da / Num. of mol.: 114 / Source method: obtained synthetically / Formula: C6H12O6 / Feature type: SUBJECT OF INVESTIGATION #10: Sugar | ChemComp-GLA / |
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-Details
Has ligand of interest | Y |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: FILAMENT / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Mastigoneme filament / Type: COMPLEX Details: Mastigoneme filaments copurified with doublet microtubules isolated from C. reinhardtii cilia Entity ID: #1-#2 / Source: NATURAL | ||||||||||||||||||||||||||||||
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Molecular weight | Experimental value: NO | ||||||||||||||||||||||||||||||
Source (natural) | Organism: Chlamydomonas reinhardtii (plant) / Strain: CC-4402 | ||||||||||||||||||||||||||||||
Buffer solution | pH: 7.4 / Details: Buffer contained protease inhibitors | ||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: Unknown concentration. Copurified with doublet microtubules. | ||||||||||||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid type: Quantifoil | ||||||||||||||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Humidity: 100 % |
-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 defocus max: 3000 nm / Nominal defocus min: 1000 nm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 39 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
-Processing
EM software | Name: PHENIX / Version: 1.21_5207: / Category: model refinement |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION |
Particle selection | Num. of particles selected: 735338 |
Symmetry | Point symmetry: C1 (asymmetric) |
3D reconstruction | Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 687452 / Algorithm: FOURIER SPACE / Symmetry type: POINT |
Atomic model building | Space: REAL |
Atomic model building | Source name: AlphaFold / Type: in silico model |