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- EMDB-35206: The cryo-EM structure of OsCyc1 hexamer state -

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Basic information

Entry
Database: EMDB / ID: EMD-35206
TitleThe cryo-EM structure of OsCyc1 hexamer state
Map data
Sample
  • Complex: Hexamer state of OsCyc1
    • Protein or peptide: Syn-copalyl diphosphate synthase, chloroplastic
Keywordssyn-copalyl diphosphate synthase / labdane-related diterpenoids / hexamer cryo-EM structure / plant defense / PLANT PROTEIN / ISOMERASE
Function / homology
Function and homology information


syn-copalyl-diphosphate synthase / syn-copalyl diphosphate synthase activity / gibberellin biosynthetic process / terpene synthase activity / chloroplast / defense response / magnesium ion binding
Similarity search - Function
: / Terpene synthase, N-terminal domain / Terpene synthase, N-terminal domain superfamily / Terpene synthase, N-terminal domain / Terpenoid cyclases/protein prenyltransferase alpha-alpha toroid / Isoprenoid synthase domain superfamily
Similarity search - Domain/homology
Syn-copalyl diphosphate synthase, chloroplastic
Similarity search - Component
Biological speciesOryza sativa Japonica Group (Japanese rice)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsMa XL / Xu HF / Tong YR / Luo YF / Dong QH / Jiang T
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Commun Chem / Year: 2023
Title: Structural and functional investigations of syn-copalyl diphosphate synthase from Oryza sativa.
Authors: Xiaoli Ma / Haifeng Xu / Yuru Tong / Yunfeng Luo / Qinghua Dong / Tao Jiang /
Abstract: The large superfamily of labdane-related diterpenoids is defined by the cyclization of linear geranylgeranyl pyrophosphate (GGPP), catalyzed by copalyl diphosphate synthases (CPSs) to form the basic ...The large superfamily of labdane-related diterpenoids is defined by the cyclization of linear geranylgeranyl pyrophosphate (GGPP), catalyzed by copalyl diphosphate synthases (CPSs) to form the basic decalin core, the copalyl diphosphates (CPPs). Three stereochemically distinct CPPs have been found in plants, namely (+)-CPP, ent-CPP and syn-CPP. Here, we used X-ray crystallography and cryo-EM methods to describe different oligomeric structures of a syn-copalyl diphosphate synthase from Oryza sativa (OsCyc1), and provided a cryo-EM structure of OsCyc1 mutant in complex with the substrate GGPP. Further analysis showed that tetramers are the dominant form of OsCyc1 in solution and are not necessary for enzyme activity in vitro. Through rational design, we identified an OsCyc1 mutant that can generate ent-CPP in addition to syn-CPP. Our work provides a structural and mechanistic basis for comparing different CPSs and paves the way for further enzyme design to obtain diterpene derivatives with specific chirality.
History
DepositionJan 29, 2023-
Header (metadata) releaseDec 6, 2023-
Map releaseDec 6, 2023-
UpdateDec 6, 2023-
Current statusDec 6, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_35206.png

Downloads & links

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Map

FileDownload / File: emd_35206.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 280 pix.
= 299.6 Å		1.07 Å/pix.
x 280 pix.
= 299.6 Å		1.07 Å/pix.
x 280 pix.
= 299.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.07 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0176
Minimum - Maximum-0.04081132 - 0.07179812
Average (Standard dev.)0.00016029603 (±0.004504701)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 299.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Hexamer state of OsCyc1

EntireName: Hexamer state of OsCyc1
Components
  • Complex: Hexamer state of OsCyc1
    • Protein or peptide: Syn-copalyl diphosphate synthase, chloroplastic

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Supramolecule #1: Hexamer state of OsCyc1

SupramoleculeName: Hexamer state of OsCyc1 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Oryza sativa Japonica Group (Japanese rice)

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Macromolecule #1: Syn-copalyl diphosphate synthase, chloroplastic

MacromoleculeName: Syn-copalyl diphosphate synthase, chloroplastic / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO / EC number: syn-copalyl-diphosphate synthase
Source (natural)Organism: Oryza sativa Japonica Group (Japanese rice)
Molecular weightTheoretical: 88.427445 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MPVFTASFQC VTLFGQPASA ADAQPLLQGQ RPFLHLHARR RRPCGPMLIS KSPPYPASEE TREWEADGQH EHTDELRETT TTMIDGIRT ALRSIGEGEI SISAYDTSLV ALLKRLDGGD GPQFPSTIDW IVQNQLPDGS WGDASFFMMG DRIMSTLACV V ALKSWNIH ...String:
MPVFTASFQC VTLFGQPASA ADAQPLLQGQ RPFLHLHARR RRPCGPMLIS KSPPYPASEE TREWEADGQH EHTDELRETT TTMIDGIRT ALRSIGEGEI SISAYDTSLV ALLKRLDGGD GPQFPSTIDW IVQNQLPDGS WGDASFFMMG DRIMSTLACV V ALKSWNIH TDKCERGLLF IQENMWRLAH EEEDWMLVGF EIALPSLLDM AKDLDLDIPY DEPALKAIYA ERERKLAKIP RD VLHSMPT TLLHSLEGMV DLDWEKLLKL RCLDGSFHCS PASTATAFQQ TGDQKCFEYL DGIVKKFNGG VPCIYPLDVY ERL WAVDRL TRLGISRHFT SEIEDCLDYI FRNWTPDGLA HTKNCPVKDI DDTAMGFRLL RLYGYQVDPC VLKKFEKDGK FFCL HGESN PSSVTPMYNT YRASQLKFPG DDGVLGRAEV FCRSFLQDRR GSNRMKDKWA IAKDIPGEVE YAMDYPWKAS LPRIE TRLY LDQYGGSGDV WIGKVLHRMT LFCNDLYLKA AKADFSNFQK ECRVELNGLR RWYLRSNLEK FGGTDPQTTL MTSYFL ASA NIFEANRAAE RLGWARVALL ADAVSSHFRR IGGPKNSTSN LEELISLVPF DDAYSGSLRE AWKQWLMAWT AKESSQE SI EGDTAILLVR AIEIFGGRHV LTGQRPDLWE YSQLEQLTSS ICCKLSRRVL AQENGESTEK VEEIDQQVDL EMQELTRR V LQGCSAINRL TRETFLHVVK SFCYVAYCSP ETIDSHIDKV IFQDVIEFHH HHHH

UniProtKB: Syn-copalyl diphosphate synthase, chloroplastic

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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

Concentration1.8 mg/mL
BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.3000000000000003 µm / Nominal defocus min: 1.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 97919
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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