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Open data
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Basic information
Entry | Database: PDB / ID: 8uc1 | ||||||
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Title | Cryo-EM structure of dolphin Prestin in low Cl buffer | ||||||
![]() | Prestin | ||||||
![]() | MEMBRANE PROTEIN / Solute Carrier / Electromotility / Voltage Sensitive / Mechanosensitive | ||||||
Function / homology | ![]() cochlear outer hair cell electromotile response / secondary active sulfate transmembrane transporter activity / oxalate transmembrane transporter activity / bicarbonate transmembrane transporter activity / chloride transmembrane transporter activity / sensory perception of sound / regulation of cell shape / plasma membrane Similarity search - Function | ||||||
Biological species | ![]() ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||
![]() | Haller, P. / Bavi, N. / Perozo, E. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Folding of prestin's anion-binding site and the mechanism of outer hair cell electromotility. Authors: Xiaoxuan Lin / Patrick R Haller / Navid Bavi / Nabil Faruk / Eduardo Perozo / Tobin R Sosnick / ![]() Abstract: Prestin responds to transmembrane voltage fluctuations by changing its cross-sectional area, a process underlying the electromotility of outer hair cells and cochlear amplification. Prestin belongs ...Prestin responds to transmembrane voltage fluctuations by changing its cross-sectional area, a process underlying the electromotility of outer hair cells and cochlear amplification. Prestin belongs to the SLC26 family of anion transporters yet is the only member capable of displaying electromotility. Prestin's voltage-dependent conformational changes are driven by the putative displacement of residue R399 and a set of sparse charged residues within the transmembrane domain, following the binding of a Cl anion at a conserved binding site formed by the amino termini of the TM3 and TM10 helices. However, a major conundrum arises as to how an anion that binds in proximity to a positive charge (R399), can promote the voltage sensitivity of prestin. Using hydrogen-deuterium exchange mass spectrometry, we find that prestin displays an unstable anion-binding site, where folding of the amino termini of TM3 and TM10 is coupled to Cl binding. This event shortens the TM3-TM10 electrostatic gap, thereby connecting the two helices, resulting in reduced cross-sectional area. These folding events upon anion binding are absent in SLC26A9, a non-electromotile transporter closely related to prestin. Dynamics of prestin embedded in a lipid bilayer closely match that in detergent micelle, except for a destabilized lipid-facing helix TM6 that is critical to prestin's mechanical expansion. We observe helix fraying at prestin's anion-binding site but cooperative unfolding of multiple lipid-facing helices, features that may promote prestin's fast electromechanical rearrangements. These results highlight a novel role of the folding equilibrium of the anion-binding site, and help define prestin's unique voltage-sensing mechanism and electromotility. | ||||||
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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PDBx/mmCIF format | ![]() | 230.6 KB | Display | ![]() |
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PDB format | ![]() | 186 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 42112MC 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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1 |
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Components
#1: Protein | Mass: 80973.750 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: SLC26A5 / Production host: ![]() #2: Chemical | Has ligand of interest | Y | |
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-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: Dolphin Prestin solubilized in GDN in low Cl buffer / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT | |||||||||||||||
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Molecular weight | Units: KILODALTONS/NANOMETER / Experimental value: NO | |||||||||||||||
Source (natural) | Organism: ![]() ![]() | |||||||||||||||
Source (recombinant) | Organism: ![]() | |||||||||||||||
Buffer solution | pH: 7.5 Details: 190 mM HEPES, 95 mM Tris-base, 1mM NaCl, 3mM DTT, 1mM EDTA, 0.02 % GDN | |||||||||||||||
Buffer component |
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Specimen | Conc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3 | |||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K / Details: 3.5s Blot time, Blot force 1 |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: TFS KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 81000 X / Nominal defocus max: 2100 nm / Nominal defocus min: 700 nm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 1.2 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
Image scans | Width: 5760 / Height: 4092 |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||
Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 170000 / Symmetry type: POINT | ||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL | ||||||||||||
Atomic model building | PDB-ID: 7S8X Accession code: 7S8X / Source name: PDB / Type: experimental model |