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- PDB-8vqj: CryoEM structure of DPOR under turnover -

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Basic information

Entry
Database: PDB / ID: 8vqj
TitleCryoEM structure of DPOR under turnover
Components
  • (Light-independent protochlorophyllide reductase subunit ...) x 2
  • Light-independent protochlorophyllide reductase iron-sulfur ATP-binding protein
KeywordsOXIDOREDUCTASE / Plant Protein / Electron Transfer Enzymes / Photosynthesis
Function / homology
Function and homology information


ferredoxin:protochlorophyllide reductase (ATP-dependent) / photosynthesis, dark reaction / light-independent bacteriochlorophyll biosynthetic process / oxidoreductase activity, acting on iron-sulfur proteins as donors / oxidoreductase activity, acting on the CH-CH group of donors, iron-sulfur protein as acceptor / 4 iron, 4 sulfur cluster binding / ATP binding / metal ion binding
Similarity search - Function
Light-independent protochlorophyllide reductase, iron-sulphur ATP-binding protein / Light-independent protochlorophyllide reductase, N subunit / : / Light-independent protochlorophyllide reductase, B subunit / Protochlorophyllide reductase, ChlB, light independent / Light-independent protochlorophyllide reductase subunit B-like, C-terminal / Proto-chlorophyllide reductase, C-terminal / Proto-chlorophyllide reductase 57 kD subunit / NifH/frxC family / NifH/chlL conserved site ...Light-independent protochlorophyllide reductase, iron-sulphur ATP-binding protein / Light-independent protochlorophyllide reductase, N subunit / : / Light-independent protochlorophyllide reductase, B subunit / Protochlorophyllide reductase, ChlB, light independent / Light-independent protochlorophyllide reductase subunit B-like, C-terminal / Proto-chlorophyllide reductase, C-terminal / Proto-chlorophyllide reductase 57 kD subunit / NifH/frxC family / NifH/chlL conserved site / 4Fe-4S iron sulfur cluster binding proteins, NifH/frxC family / NifH/frxC family signature 2. / NifH/frxC family signature 1. / NIFH_FRXC family profile. / : / Nitrogenase/oxidoreductase, component 1 / Nitrogenase component 1 type Oxidoreductase / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
COPPER (II) ION / Protochlorophyllide / IRON/SULFUR CLUSTER / Light-independent protochlorophyllide reductase subunit N / Light-independent protochlorophyllide reductase subunit B / Light-independent protochlorophyllide reductase iron-sulfur ATP-binding protein
Similarity search - Component
Biological speciesCereibacter sphaeroides (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.82 Å
AuthorsKashyap, R. / Antony, E.
Funding support United States, 1items
OrganizationGrant numberCountry
Department of Energy (DOE, United States)DE-SC0020965 United States
CitationJournal: Nat Commun / Year: 2025
Title: Cryo-EM captures the coordination of asymmetric electron transfer through a di-copper site in DPOR.
Authors: Rajnandani Kashyap / Natalie Walsh / Jaigeeth Deveryshetty / Monika Tokmina-Lukaszewska / Kewei Zhao / Yunqiao J Gan / Brian M Hoffman / Ritimukta Sarangi / Brian Bothner / Brian Bennett / Edwin Antony /
Abstract: Enzymes that catalyze long-range electron transfer (ET) reactions often function as higher order complexes that possess two structurally symmetrical halves. The functional advantages for such an ...Enzymes that catalyze long-range electron transfer (ET) reactions often function as higher order complexes that possess two structurally symmetrical halves. The functional advantages for such an architecture remain a mystery. Using cryoelectron microscopy we capture snapshots of the nitrogenase-like dark-operative protochlorophyllide oxidoreductase (DPOR) during substrate binding and turnover. DPOR catalyzes reduction of the C17 = C18 double bond in protochlorophyllide during the dark chlorophyll biosynthetic pathway. DPOR is composed of electron donor (L-protein) and acceptor (NB-protein) component proteins that transiently form a complex in the presence of ATP to facilitate ET. NB-protein is an αβ heterotetramer with two structurally identical halves. However, our structures reveal that NB-protein becomes functionally asymmetric upon substrate binding. Asymmetry results in allosteric inhibition of L-protein engagement and ET in one half. Residues that form a conduit for ET are aligned in one half while misaligned in the other. An ATP hydrolysis-coupled conformational switch is triggered once ET is accomplished in one half. These structural changes are then relayed to the other half through a di-nuclear copper center at the tetrameric interface of the NB-protein and leads to activation of ET and substrate reduction. These findings provide a mechanistic blueprint for regulation of long-range electron transfer reactions.
History
DepositionJan 18, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 30, 2025Provider: repository / Type: Initial release
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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Light-independent protochlorophyllide reductase subunit N
B: Light-independent protochlorophyllide reductase subunit B
C: Light-independent protochlorophyllide reductase subunit N
D: Light-independent protochlorophyllide reductase subunit B
E: Light-independent protochlorophyllide reductase iron-sulfur ATP-binding protein
F: Light-independent protochlorophyllide reductase iron-sulfur ATP-binding protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)281,28315
Polymers278,8276
Non-polymers2,4579
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Light-independent protochlorophyllide reductase subunit ... , 2 types, 4 molecules ACBD

#1: Protein Light-independent protochlorophyllide reductase subunit N / DPOR subunit N / LI-POR subunit N


Mass: 46188.773 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Cereibacter sphaeroides (bacteria) / Gene: bchN, RSKD131_1611 / Production host: Escherichia coli (E. coli)
References: UniProt: B9KK24, ferredoxin:protochlorophyllide reductase (ATP-dependent)
#2: Protein Light-independent protochlorophyllide reductase subunit B / DPOR subunit B / LI-POR subunit B


Mass: 58374.266 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Cereibacter sphaeroides (bacteria) / Gene: bchB, RSKD131_1612 / Production host: Escherichia coli (E. coli)
References: UniProt: B9KK25, ferredoxin:protochlorophyllide reductase (ATP-dependent)

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Protein , 1 types, 2 molecules EF

#3: Protein Light-independent protochlorophyllide reductase iron-sulfur ATP-binding protein / LI-POR subunit L


Mass: 34850.340 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Cereibacter sphaeroides (bacteria) / Gene: bchL, RHOS4_18930, RSP_0288 / Production host: Escherichia coli (E. coli)
References: UniProt: Q9RFD6, ferredoxin:protochlorophyllide reductase (ATP-dependent)

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Non-polymers , 4 types, 9 molecules

#4: Chemical ChemComp-SF4 / IRON/SULFUR CLUSTER


Mass: 351.640 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Fe4S4 / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical ChemComp-PMR / Protochlorophyllide


Mass: 612.957 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C35H32MgN4O5 / Feature type: SUBJECT OF INVESTIGATION
#6: Chemical ChemComp-CU / COPPER (II) ION


Mass: 63.546 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Cu / Feature type: SUBJECT OF INVESTIGATION
#7: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY
Has protein modificationN

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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-IDSource
1CryoEM structure of DPOR under turnoverCOMPLEX#1-#30RECOMBINANT
2CryoEM structure of DPOR under turnoverCOMPLEX#1-#31RECOMBINANT
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
110.3078 MDaNO
210.236 MDaNO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Cereibacter sphaeroides (bacteria)1063
22Cereibacter sphaeroides (bacteria)1063
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDStrain
21Escherichia coli (E. coli)562BL21(DE3)Suf++ strain (PK11466)
22Escherichia coli (E. coli)562
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal magnification: 105000 X / Nominal defocus max: 2200 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM softwareName: PHENIX / Category: model refinement
CTF correctionType: NONE
3D reconstructionResolution: 3.82 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 18965 / Symmetry type: POINT
RefinementHighest resolution: 3.82 Å
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00917498
ELECTRON MICROSCOPYf_angle_d1.88323854
ELECTRON MICROSCOPYf_dihedral_angle_d11.442501
ELECTRON MICROSCOPYf_chiral_restr0.3452694
ELECTRON MICROSCOPYf_plane_restr0.0063107

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