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- PDB-8a9b: Single Particle cryo-EM of the empty lipid binding protein P116 (... -

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Basic information

Entry
Database: PDB / ID: 8a9b
TitleSingle Particle cryo-EM of the empty lipid binding protein P116 (MPN213) from Mycoplasma pneumoniae at 4 Angstrom resolution
ComponentsLipid binding protein P116 (MPN213)
KeywordsLIPID BINDING PROTEIN / Empty lipid binding protein P116 (MPN213) from Mycoplasma pneumoniae / LIPID BINDING
Function / homologymembrane / Uncharacterized protein MG075 homolog
Function and homology information
Biological speciesMycoplasma pneumoniae M129 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4 Å
AuthorsSprankel, L. / Vizarraga, D.
Funding support Germany, 3items
OrganizationGrant numberCountry
German Research Foundation (DFG)FR 1653/14-1 Germany
German Research Foundation (DFG)FR 1653/6-3 Germany
German Research Foundation (DFG)GRK 2566/1 Germany
CitationJournal: Nat Struct Mol Biol / Year: 2023
Title: Essential protein P116 extracts cholesterol and other indispensable lipids for Mycoplasmas.
Authors: Lasse Sprankel / David Vizarraga / Jesús Martín / Sina Manger / Jakob Meier-Credo / Marina Marcos / Josep Julve / Noemi Rotllan / Margot P Scheffer / Joan Carles Escolà-Gil / Julian D ...Authors: Lasse Sprankel / David Vizarraga / Jesús Martín / Sina Manger / Jakob Meier-Credo / Marina Marcos / Josep Julve / Noemi Rotllan / Margot P Scheffer / Joan Carles Escolà-Gil / Julian D Langer / Jaume Piñol / Ignacio Fita / Achilleas S Frangakis /
Abstract: Mycoplasma pneumoniae, responsible for approximately 30% of community-acquired human pneumonia, needs to extract lipids from the host environment for survival and proliferation. Here, we report a ...Mycoplasma pneumoniae, responsible for approximately 30% of community-acquired human pneumonia, needs to extract lipids from the host environment for survival and proliferation. Here, we report a comprehensive structural and functional analysis of the previously uncharacterized protein P116 (MPN_213). Single-particle cryo-electron microscopy of P116 reveals a homodimer presenting a previously unseen fold, forming a huge hydrophobic cavity, which is fully accessible to solvent. Lipidomics analysis shows that P116 specifically extracts lipids such as phosphatidylcholine, sphingomyelin and cholesterol. Structures of different conformational states reveal the mechanism by which lipids are extracted. This finding immediately suggests a way to control Mycoplasma infection by interfering with lipid uptake.
History
DepositionJun 28, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 22, 2023Provider: repository / Type: Initial release
Revision 1.1Mar 29, 2023Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: Lipid binding protein P116 (MPN213)


Theoretical massNumber of molelcules
Total (without water)114,1501
Polymers114,1501
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Lipid binding protein P116 (MPN213) / Uncharacterized protein MG075 homolog


Mass: 114149.852 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mycoplasma pneumoniae M129 (bacteria) / Strain: ATCC 29342 / M129 / Gene: MPN_213, G07_orf1030, MP618 / Production host: Escherichia coli B83 (bacteria) / References: UniProt: P75556

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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: Empty P116 monomer / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Mycoplasma pneumoniae M129 (bacteria)
Source (recombinant)Organism: Escherichia coli B83 (bacteria)
Buffer solutionpH: 7.4
Buffer componentConc.: 20 mM / Name: Tris
SpecimenConc.: 0.3 mg/ml / Embedding 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: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 130000 X / Calibrated magnification: 130000 X / Nominal defocus max: 3500 nm / Nominal defocus min: 1000 nm / Calibrated defocus min: 1000 nm / Calibrated defocus max: 3500 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: BASIC
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 15299
EM imaging opticsEnergyfilter name: GIF Quantum SE / Energyfilter slit width: 20 eV
Image scansMovie frames/image: 34

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM software
IDNameVersionCategory
1cryoSPARC3.3.2particle selection
4cryoSPARC3.32CTF correction
13cryoSPARC3.3.23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 4532601
3D reconstructionResolution: 4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 633332 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0046495
ELECTRON MICROSCOPYf_angle_d0.8678750
ELECTRON MICROSCOPYf_dihedral_angle_d6.409838
ELECTRON MICROSCOPYf_chiral_restr0.05999
ELECTRON MICROSCOPYf_plane_restr0.0051119

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