+Open data
-Basic information
Entry | Database: PDB / ID: 8j5z | ||||||
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Title | The cryo-EM structure of the TwOSC1 tetramer | ||||||
Components | Terpene cyclase/mutase family member | ||||||
Keywords | PLANT PROTEIN / oxidosqualene cyclase / Tripterygium wilfordii Hook. f. / TwOSC1 / friedelin / cryo-EM structure | ||||||
Function / homology | Function and homology information oxidosqualene cyclase activity / triterpenoid biosynthetic process / Isomerases; Intramolecular transferases; Transferring other groups / lipid droplet Similarity search - Function | ||||||
Biological species | Tripterygium wilfordii (plant) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.75 Å | ||||||
Authors | Ma, X. / Yuru, T. / Yunfeng, L. / Jiang, T. | ||||||
Funding support | 1items
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Citation | Journal: Angew Chem Int Ed Engl / Year: 2023 Title: Structural and Catalytic Insight into the Unique Pentacyclic Triterpene Synthase TwOSC. Authors: Yunfeng Luo / Xiaoli Ma / Yufan Qiu / Yun Lu / Siyu Shen / Yang Li / Haiyun Gao / Kang Chen / Jiawei Zhou / Tianyuan Hu / Lichan Tu / Huan Zhao / Dan Li / Faqiang Leng / Wei Gao / Tao Jiang ...Authors: Yunfeng Luo / Xiaoli Ma / Yufan Qiu / Yun Lu / Siyu Shen / Yang Li / Haiyun Gao / Kang Chen / Jiawei Zhou / Tianyuan Hu / Lichan Tu / Huan Zhao / Dan Li / Faqiang Leng / Wei Gao / Tao Jiang / Changli Liu / Luqi Huang / Ruibo Wu / Yuru Tong / Abstract: The oxidosqualene cyclase (OSC) catalyzed cyclization of the linear substrate (3S)-2,3-oxidosqualene to form diverse pentacyclic triterpenoid (PT) skeletons is one of the most complex reactions in ...The oxidosqualene cyclase (OSC) catalyzed cyclization of the linear substrate (3S)-2,3-oxidosqualene to form diverse pentacyclic triterpenoid (PT) skeletons is one of the most complex reactions in nature. Friedelin has a unique PT skeleton involving a fascinating nine-step cation shuttle run (CSR) cascade rearrangement reaction, in which the carbocation formed at C2 moves to the other side of the skeleton, runs back to C3 to yield a friedelin cation, which is finally deprotonated. However, as crystal structure data of plant OSCs are lacking, it remains unknown why the CSR cascade reactions occur in friedelin biosynthesis, as does the exact catalytic mechanism of the CSR. In this study, we determined the first cryogenic electron microscopy structure of a plant OSC, friedelin synthase, from Tripterygium wilfordii Hook. f (TwOSC). We also performed quantum mechanics/molecular mechanics simulations to reveal the energy profile for the CSR cascade reaction and identify key residues crucial for PT skeleton formation. Furthermore, we semirationally designed two TwOSC mutants, which significantly improved the yields of friedelin and β-amyrin, respectively. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8j5z.cif.gz | 537.1 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8j5z.ent.gz | 442.6 KB | Display | PDB format |
PDBx/mmJSON format | 8j5z.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8j5z_validation.pdf.gz | 1.4 MB | Display | wwPDB validaton report |
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Full document | 8j5z_full_validation.pdf.gz | 1.4 MB | Display | |
Data in XML | 8j5z_validation.xml.gz | 86.2 KB | Display | |
Data in CIF | 8j5z_validation.cif.gz | 124.8 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/j5/8j5z ftp://data.pdbj.org/pub/pdb/validation_reports/j5/8j5z | HTTPS FTP |
-Related structure data
Related structure data | 35996MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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Noncrystallographic symmetry (NCS) | NCS domain:
NCS domain segments: Ens-ID: ens_1
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-Components
#1: Protein | Mass: 88212.500 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Tripterygium wilfordii (plant) / Gene: OSC1 / Production host: Komagataella pastoris (fungus) / References: UniProt: A0A499QXI7 #2: Sugar | ChemComp-BOG / Has ligand of interest | Y | |
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-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: Tetramer of TwOSC1 / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT |
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Source (natural) | Organism: Tripterygium wilfordii (plant) |
Source (recombinant) | Organism: Komagataella pastoris (fungus) |
Buffer solution | pH: 8 Details: 50 mM Tris-HCl (pH 8.0), 300 mM NaCl, 2 mM dithiothreitol, and 0.2% OG |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-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: 2300 nm / Nominal defocus min: 1500 nm |
Image recording | Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
Software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.75 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 32698 / Symmetry type: POINT | ||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE | ||||||||||||||||||||||||||||
Refine LS restraints |
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Refine LS restraints NCS |
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