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- PDB-8p4x: FAD_ox bound dark state structure of PdLCry -

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Basic information

Entry
Database: PDB / ID: 8p4x
TitleFAD_ox bound dark state structure of PdLCry
ComponentsPutative light-receptive cryptochrome (Fragment)
KeywordsFLAVOPROTEIN / light-sensitive / circalunar clock
Function / homology
Function and homology information


entrainment of circadian clock by photoperiod / FAD binding / circadian regulation of gene expression / negative regulation of DNA-templated transcription / perinuclear region of cytoplasm / DNA binding / nucleus
Similarity search - Function
Cryptochrome/DNA photolyase class 1 / Cryptochrome/DNA photolyase, FAD-binding domain / FAD binding domain of DNA photolyase / DNA photolyase, N-terminal / Cryptochrome/photolyase, N-terminal domain superfamily / DNA photolyase / Photolyase/cryptochrome alpha/beta domain profile. / Cryptochrome/DNA photolyase, FAD-binding domain-like superfamily / Rossmann-like alpha/beta/alpha sandwich fold
Similarity search - Domain/homology
FLAVIN-ADENINE DINUCLEOTIDE / Cryptochrome-1
Similarity search - Component
Biological speciesPlatynereis dumerilii (Dumeril's clam worm)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.57 Å
AuthorsBehrmann, E. / Behrmann, H.
Funding support Germany, 4items
OrganizationGrant numberCountry
German Research Foundation (DFG)INST 216/949-1 FUGG Germany
German Research Foundation (DFG)INST 216/512/1 FUGG Germany
German Research Foundation (DFG)SFB1372 Germany
German Research Foundation (DFG)GRK1885 Germany
CitationJournal: Nat Commun / Year: 2023
Title: A marine cryptochrome with an inverse photo-oligomerization mechanism.
Authors: Hong Ha Vu / Heide Behrmann / Maja Hanić / Gayathri Jeyasankar / Shruthi Krishnan / Dennis Dannecker / Constantin Hammer / Monika Gunkel / Ilia A Solov'yov / Eva Wolf / Elmar Behrmann /
Abstract: Cryptochromes (CRYs) are a structurally conserved but functionally diverse family of proteins that can confer unique sensory properties to organisms. In the marine bristle worm Platynereis dumerilii, ...Cryptochromes (CRYs) are a structurally conserved but functionally diverse family of proteins that can confer unique sensory properties to organisms. In the marine bristle worm Platynereis dumerilii, its light receptive cryptochrome L-CRY (PdLCry) allows the animal to discriminate between sunlight and moonlight, an important requirement for synchronizing its lunar cycle-dependent mass spawning. Using cryo-electron microscopy, we show that in the dark, PdLCry adopts a dimer arrangement observed neither in plant nor insect CRYs. Intense illumination disassembles the dimer into monomers. Structural and functional data suggest a mechanistic coupling between the light-sensing flavin adenine dinucleotide chromophore, the dimer interface, and the C-terminal tail helix, with a likely involvement of the phosphate binding loop. Taken together, our work establishes PdLCry as a CRY protein with inverse photo-oligomerization with respect to plant CRYs, and provides molecular insights into how this protein might help discriminating the different light intensities associated with sunlight and moonlight.
History
DepositionMay 23, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 8, 2023Provider: repository / Type: Initial release
Revision 1.1Mar 27, 2024Group: Database references / Category: citation
Item: _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Putative light-receptive cryptochrome (Fragment)
B: Putative light-receptive cryptochrome (Fragment)
hetero molecules


Theoretical massNumber of molelcules
Total (without water)132,5856
Polymers130,9662
Non-polymers1,6204
Water1,51384
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1chain "B"
d_2ens_1chain "A"

NCS domain segments:

Ens-ID: ens_1

Dom-IDComponent-IDBeg auth comp-IDBeg label comp-IDEnd auth comp-IDEnd label comp-IDAuth asym-IDLabel asym-IDAuth seq-IDLabel seq-ID
d_11GLUGLUCYSCYSBB31 - 55631 - 556
d_12FADFADFADFADBE601
d_21GLUGLUCYSCYSAA31 - 55631 - 556
d_22FADFADFADFADAC601

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Components

#1: Protein Putative light-receptive cryptochrome (Fragment)


Mass: 65482.855 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Platynereis dumerilii (Dumeril's clam worm)
Plasmid: pCoofy27 / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: G8Z9G7
#2: Chemical ChemComp-FAD / FLAVIN-ADENINE DINUCLEOTIDE


Mass: 785.550 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Formula: C27H33N9O15P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: FAD*YM
#3: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 84 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY

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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

ComponentName: dimeric complex of PdLCry in the dark state / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.13 MDa / Experimental value: YES
Source (natural)Organism: Platynereis dumerilii (Dumeril's clam worm)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm) / Strain: Sf9
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
125 mMBis-tris propane1
2150 mMNaCl1
31 mMTCEP1
40.01 %fOM1
SpecimenConc.: 0.7 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: grids were prepared under far-red light illumination (Osram OSLON SSL 120, GF CSSPM1.24)
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 293.15 K
Details: grids were prepared under far-red light illumination

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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 FIELD / Nominal magnification: 96000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 300 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 30 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k) / Details: see material+methods

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Processing

EM software
IDNameVersionCategory
1cryoSPARCparticle selection
2EPUimage acquisition
4cryoSPARCCTF correction
7Cootmodel fitting
8PHENIXmodel fitting
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
13cryoSPARC3D reconstruction
14PHENIX1.20.1_4487:model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionDetails: see supplementary table and materials+methods
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 2.57 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 424744 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: REAL
Atomic model buildingDetails: starting model was based on 4jzy / Source name: Other / Type: integrative model
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 22.58 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0038964
ELECTRON MICROSCOPYf_angle_d0.47312194
ELECTRON MICROSCOPYf_dihedral_angle_d11.9623276
ELECTRON MICROSCOPYf_chiral_restr0.0391220
ELECTRON MICROSCOPYf_plane_restr0.0041552
Refine LS restraints NCSType: NCS constraints / Rms dev position: 2.57425599462E-13 Å

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