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- PDB-9bkv: DosP R97A bent form -

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Basic information

Entry
Database: PDB / ID: 9bkv
TitleDosP R97A bent form
ComponentsOxygen sensor protein DosP
KeywordsOXYGEN BINDING / Heme / DosP / Phosphodiesterase / c-di-GMP
Function / homology
Function and homology information


cyclic-guanylate-specific phosphodiesterase / cyclic-guanylate-specific phosphodiesterase activity / response to oxygen levels / oxygen sensor activity / heme binding / magnesium ion binding / protein homodimerization activity
Similarity search - Function
Diguanylate cyclase/phosphodiesterase / : / Putative diguanylate phosphodiesterase / EAL domain / EAL domain superfamily / EAL domain profile. / EAL domain / Diguanylate cyclase, GGDEF domain / diguanylate cyclase / GGDEF domain profile. ...Diguanylate cyclase/phosphodiesterase / : / Putative diguanylate phosphodiesterase / EAL domain / EAL domain superfamily / EAL domain profile. / EAL domain / Diguanylate cyclase, GGDEF domain / diguanylate cyclase / GGDEF domain profile. / GGDEF domain / PAS domain / PAS-associated, C-terminal / PAC domain profile. / Nucleotide cyclase / PAC motif / Motif C-terminal to PAS motifs (likely to contribute to PAS structural domain) / PAS domain / PAS repeat profile. / PAS domain / PAS domain superfamily / Reverse transcriptase/Diguanylate cyclase domain
Similarity search - Domain/homology
PROTOPORPHYRIN IX CONTAINING FE / Oxygen sensor protein DosP
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.43 Å
AuthorsKumar, P. / Kober, D.L.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM141261 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM129547 United States
CitationJournal: Nat Commun / Year: 2024
Title: Structures of the multi-domain oxygen sensor DosP: remote control of a c-di-GMP phosphodiesterase by a regulatory PAS domain.
Authors: Wenbi Wu / Pankaj Kumar / Chad A Brautigam / Shih-Chia Tso / Hamid R Baniasadi / Daniel L Kober / Marie-Alda Gilles-Gonzalez /
Abstract: The heme-based direct oxygen sensor DosP degrades c-di-GMP, a second messenger nearly unique to bacteria. In stationary phase Escherichia coli, DosP is the most abundant c-di-GMP phosphodiesterase. ...The heme-based direct oxygen sensor DosP degrades c-di-GMP, a second messenger nearly unique to bacteria. In stationary phase Escherichia coli, DosP is the most abundant c-di-GMP phosphodiesterase. Ligation of O to a heme-binding PAS domain (hPAS) of the protein enhances the phosphodiesterase through an allosteric mechanism that has remained elusive. We determine six structures of full-length DosP in its aerobic or anaerobic conformations, with or without c-di-GMP. DosP is an elongated dimer with the regulatory heme containing domain and phosphodiesterase separated by nearly 180 Å. In the absence of substrate, regardless of the heme status, DosP presents an equilibrium of two distinct conformations. Binding of substrate induces DosP to adopt a single, ON-state or OFF-state conformation depending on its heme status. Structural and biochemical studies of this multi-domain sensor and its mutants provide insights into signal regulation of second-messenger levels.
History
DepositionApr 29, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 20, 2024Provider: repository / Type: Initial release
Revision 1.1Nov 27, 2024Group: Data collection / Database references / Category: citation / em_admin
Item: _citation.page_last / _citation.pdbx_database_id_PubMed ..._citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Oxygen sensor protein DosP
B: Oxygen sensor protein DosP
hetero molecules


Theoretical massNumber of molelcules
Total (without water)183,5954
Polymers182,3622
Non-polymers1,2332
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Oxygen sensor protein DosP / Direct oxygen-sensing phosphodiesterase / Direct oxygen sensor protein / Ec DOS / Heme-regulated ...Direct oxygen-sensing phosphodiesterase / Direct oxygen sensor protein / Ec DOS / Heme-regulated cyclic di-GMP phosphodiesterase


Mass: 91181.156 Da / Num. of mol.: 2 / Mutation: R97A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: dosP, dos, pdeO, yddU, b1489, JW1484 / Production host: Escherichia coli (E. coli)
References: UniProt: P76129, cyclic-guanylate-specific phosphodiesterase
#2: Chemical ChemComp-HEM / PROTOPORPHYRIN IX CONTAINING FE / HEME


Mass: 616.487 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C34H32FeN4O4 / Feature type: SUBJECT OF INVESTIGATION
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

ComponentName: Dimer of DosP / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.2 MDa / Experimental value: NO
Source (natural)Organism: Escherichia coli (E. coli)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationCryogen name: ETHANE

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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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM software
IDNameVersionCategory
10cryoSPARC4.2initial Euler assignment
11cryoSPARC4.2final Euler assignment
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.43 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 391512 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00612832
ELECTRON MICROSCOPYf_angle_d0.81317414
ELECTRON MICROSCOPYf_dihedral_angle_d6.2551710
ELECTRON MICROSCOPYf_chiral_restr0.0481927
ELECTRON MICROSCOPYf_plane_restr0.0052265

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