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- PDB-6jbh: Cryo-EM structure and transport mechanism of a wall teichoic acid... -

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Basic information

Entry
Database: PDB / ID: 6jbh
TitleCryo-EM structure and transport mechanism of a wall teichoic acid ABC transporter
Components
  • TarG
  • TarH
KeywordsTRANSPORT PROTEIN / ABC Transporter
Biological speciesAlicyclobacillus herbarius (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.94 Å
AuthorsChen, L. / Hou, W.T. / Fan, T. / Li, Y.H. / Liu, B.H. / Jiang, Y.L. / Sun, L.F. / Chen, Y. / Zhou, C.Z.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China2015CB910103 China
CitationJournal: mBio / Year: 2020
Title: Cryo-electron Microscopy Structure and Transport Mechanism of a Wall Teichoic Acid ABC Transporter.
Authors: Li Chen / Wen-Tao Hou / Tao Fan / Banghui Liu / Ting Pan / Yu-Hui Li / Yong-Liang Jiang / Wen Wen / Zhi-Peng Chen / Linfeng Sun / Cong-Zhao Zhou / Yuxing Chen /
Abstract: The wall teichoic acid (WTA) is a major cell wall component of Gram-positive bacteria, such as methicillin-resistant (MRSA), a common cause of fatal clinical infections in humans. Thus, the ...The wall teichoic acid (WTA) is a major cell wall component of Gram-positive bacteria, such as methicillin-resistant (MRSA), a common cause of fatal clinical infections in humans. Thus, the indispensable ABC transporter TarGH, which flips WTA from cytoplasm to extracellular space, becomes a promising target of anti-MRSA drugs. Here, we report the 3.9-Å cryo-electron microscopy (cryo-EM) structure of a 50% sequence-identical homolog of TarGH from at an ATP-free and inward-facing conformation. Structural analysis combined with activity assays enables us to clearly decode the binding site and inhibitory mechanism of the anti-MRSA inhibitor Targocil, which targets TarGH. Moreover, we propose a "crankshaft conrod" mechanism utilized by TarGH, which can be applied to similar ABC transporters that translocate a rather big substrate through relatively subtle conformational changes. These findings provide a structural basis for the rational design and optimization of antibiotics against MRSA. The wall teichoic acid (WTA) is a major component of cell wall and a pathogenic factor in methicillin-resistant (MRSA). The ABC transporter TarGH is indispensable for flipping WTA precursor from cytoplasm to the extracellular space, thus making it a promising drug target for anti-MRSA agents. The 3.9-Å cryo-EM structure of a TarGH homolog helps us to decode the binding site and inhibitory mechanism of a recently reported inhibitor, Targocil, and provides a structural platform for rational design and optimization of potential antibiotics. Moreover, we propose a "crankshaft conrod" mechanism to explain how a big substrate is translocated through subtle conformational changes of type II exporters. These findings advance our understanding of anti-MRSA drug design and ABC transporters.
History
DepositionJan 25, 2019Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Mar 4, 2020Provider: repository / Type: Initial release
Revision 1.1Apr 8, 2020Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Mar 27, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
A: TarH
B: TarH
C: TarG
D: TarG


Theoretical massNumber of molelcules
Total (without water)126,5364
Polymers126,5364
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area9640 Å2
ΔGint-53 kcal/mol
Surface area53620 Å2
MethodPISA

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Components

#1: Protein TarH


Mass: 30201.881 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Alicyclobacillus herbarius (bacteria) / Production host: Escherichia coli (E. coli)
#2: Protein TarG


Mass: 33066.285 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Alicyclobacillus herbarius (bacteria) / Production host: Escherichia coli (E. coli)
Sequence detailsSequences of TarG has deposited to NCBI with accession ID WP_026962790.1, and N-terminal ...Sequences of TarG has deposited to NCBI with accession ID WP_026962790.1, and N-terminal MGHHHHHHHHHH are from expression tag. Sequence of TarH has been deposited with accession ID WP_026962789.1.

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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: wall teichoic acid ABC transporter TarGH / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Alicyclobacillus herbarius (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenConc.: 8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 281 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
Image recordingElectron dose: 64 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.14_3228: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.94 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 95416 / Symmetry type: POINT
Atomic model buildingB value: 198.567 / Protocol: AB INITIO MODEL

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