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Open data
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Basic information
Entry | Database: PDB / ID: 8tth | ||||||||||||||||||||||||||||||
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Title | NorA single mutant - D307N at pH 7.5 | ||||||||||||||||||||||||||||||
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![]() | TRANSPORT PROTEIN/IMMUNE SYSTEM / D307N NorA / efflux pump / MRSA / TRANSPORT PROTEIN / TRANSPORT PROTEIN-IMMUNE SYSTEM complex | ||||||||||||||||||||||||||||||
Function / homology | ![]() | ||||||||||||||||||||||||||||||
Biological species | ![]() ![]() ![]() | ||||||||||||||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.54 Å | ||||||||||||||||||||||||||||||
![]() | Li, J.P. / Li, Y. / Koide, A. / Kuang, H.H. / Torres, V.J. / Koide, S. / Wang, D.N. / Traaseth, N.J. | ||||||||||||||||||||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Proton-coupled transport mechanism of the efflux pump NorA. Authors: Jianping Li / Yan Li / Akiko Koide / Huihui Kuang / Victor J Torres / Shohei Koide / Da-Neng Wang / Nathaniel J Traaseth / ![]() Abstract: Efflux pump antiporters confer drug resistance to bacteria by coupling proton import with the expulsion of antibiotics from the cytoplasm. Despite efforts there remains a lack of understanding as to ...Efflux pump antiporters confer drug resistance to bacteria by coupling proton import with the expulsion of antibiotics from the cytoplasm. Despite efforts there remains a lack of understanding as to how acid/base chemistry drives drug efflux. Here, we uncover the proton-coupling mechanism of the Staphylococcus aureus efflux pump NorA by elucidating structures in various protonation states of two essential acidic residues using cryo-EM. Protonation of Glu222 and Asp307 within the C-terminal domain stabilized the inward-occluded conformation by forming hydrogen bonds between the acidic residues and a single helix within the N-terminal domain responsible for occluding the substrate binding pocket. Remarkably, deprotonation of both Glu222 and Asp307 is needed to release interdomain tethering interactions, leading to opening of the pocket for antibiotic entry. Hence, the two acidic residues serve as a "belt and suspenders" protection mechanism to prevent simultaneous binding of protons and drug that enforce NorA coupling stoichiometry and confer antibiotic resistance. | ||||||||||||||||||||||||||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 116.1 KB | Display | ![]() |
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PDB format | ![]() | 86.5 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.2 MB | Display | ![]() |
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Full document | ![]() | 1.2 MB | Display | |
Data in XML | ![]() | 33 KB | Display | |
Data in CIF | ![]() | 47.1 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 41608MC ![]() 8tteC ![]() 8ttfC ![]() 8ttgC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 46654.863 Da / Num. of mol.: 1 / Mutation: D307N Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() |
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#2: Antibody | Mass: 14070.568 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
#3: Antibody | Mass: 11209.451 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: D307N-NorA:FabDA1 / Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES | |||||||||||||||
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Molecular weight | Value: 0.100656 MDa / Experimental value: NO | |||||||||||||||
Source (natural) | Organism: ![]() ![]() | |||||||||||||||
Source (recombinant) | Organism: ![]() ![]() | |||||||||||||||
Buffer solution | pH: 7.5 | |||||||||||||||
Buffer component |
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Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||
Vitrification | Cryogen name: ETHANE |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 105000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 77 K / Temperature (min): 77 K |
Image recording | Average exposure time: 1.2 sec. / Electron dose: 52.96 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
Image scans | Sampling size: 5 µm / Width: 5760 / Height: 4092 |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.54 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 136284 / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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