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- PDB-8otx: Cryo-EM structure of Strongylocentrotus purpuratus sperm-specific... -

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Basic information

Entry
Database: PDB / ID: 8otx
TitleCryo-EM structure of Strongylocentrotus purpuratus sperm-specific Na+/H+ exchanger SLC9C1 in nanodisc
ComponentsSperm-specific sodium proton exchanger
KeywordsTRANSPORT PROTEIN / VSD / CNBD / Transporter / Voltage sensor / exchanger / sperm motility
Function / homology
Function and homology information


potassium:proton antiporter activity / sperm head / sodium:proton antiporter activity / sodium ion import across plasma membrane / cAMP-mediated signaling / cGMP binding / sperm flagellum / cAMP binding / potassium ion transmembrane transport / regulation of intracellular pH ...potassium:proton antiporter activity / sperm head / sodium:proton antiporter activity / sodium ion import across plasma membrane / cAMP-mediated signaling / cGMP binding / sperm flagellum / cAMP binding / potassium ion transmembrane transport / regulation of intracellular pH / protein homodimerization activity / plasma membrane
Similarity search - Function
Cation/H+ exchanger, CPA1 family / Cation/H+ exchanger / Sodium/hydrogen exchanger family / Cyclic nucleotide-monophosphate binding domain / Cyclic nucleotide-binding domain / cAMP/cGMP binding motif profile. / Cyclic nucleotide-binding domain / Cyclic nucleotide-binding domain superfamily / Voltage-dependent channel domain superfamily / RmlC-like jelly roll fold
Similarity search - Domain/homology
1,2-DIACYL-GLYCEROL-3-SN-PHOSPHATE / Sperm-specific sodium:proton exchanger
Similarity search - Component
Biological speciesStrongylocentrotus purpuratus (purple sea urchin)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.08 Å
AuthorsYeo, H. / Mehta, V. / Gulati, A. / Drew, D.
Funding supportEuropean Union, 1items
OrganizationGrant numberCountry
European Research Council (ERC)ERC-CoG-820187European Union
CitationJournal: Nature / Year: 2023
Title: Structure and electromechanical coupling of a voltage-gated Na/H exchanger.
Authors: Hyunku Yeo / Ved Mehta / Ashutosh Gulati / David Drew /
Abstract: Voltage-sensing domains control the activation of voltage-gated ion channels, with a few exceptions. One such exception is the sperm-specific Na/H exchanger SLC9C1, which is the only known ...Voltage-sensing domains control the activation of voltage-gated ion channels, with a few exceptions. One such exception is the sperm-specific Na/H exchanger SLC9C1, which is the only known transporter to be regulated by voltage-sensing domains. After hyperpolarization of sperm flagella, SLC9C1 becomes active, causing pH alkalinization and CatSper Ca channel activation, which drives chemotaxis. SLC9C1 activation is further regulated by cAMP, which is produced by soluble adenyl cyclase (sAC). SLC9C1 is therefore an essential component of the pH-sAC-cAMP signalling pathway in metazoa, required for sperm motility and fertilization. Despite its importance, the molecular basis of SLC9C1 voltage activation is unclear. Here we report cryo-electron microscopy (cryo-EM) structures of sea urchin SLC9C1 in detergent and nanodiscs. We show that the voltage-sensing domains are positioned in an unusual configuration, sandwiching each side of the SLC9C1 homodimer. The S4 segment is very long, 90 Å in length, and connects the voltage-sensing domains to the cytoplasmic cyclic-nucleotide-binding domains. The S4 segment is in the up configuration-the inactive state of SLC9C1. Consistently, although a negatively charged cavity is accessible for Na to bind to the ion-transporting domains of SLC9C1, an intracellular helix connected to S4 restricts their movement. On the basis of the differences in the cryo-EM structure of SLC9C1 in the presence of cAMP, we propose that, upon hyperpolarization, the S4 segment moves down, removing this constriction and enabling Na/H exchange.
History
DepositionApr 21, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 1, 2023Provider: repository / Type: Initial release
Revision 1.1Nov 8, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID
Revision 1.2Nov 15, 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
Revision 1.3Nov 22, 2023Group: Database references / Category: citation_author / Item: _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
B: Sperm-specific sodium proton exchanger
A: Sperm-specific sodium proton exchanger
hetero molecules


Theoretical massNumber of molelcules
Total (without water)297,8836
Polymers295,0632
Non-polymers2,8204
Water362
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Sperm-specific sodium proton exchanger


Mass: 147531.328 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Strongylocentrotus purpuratus (purple sea urchin)
Production host: Homo sapiens (human) / References: UniProt: A3RL54
#2: Chemical
ChemComp-3PH / 1,2-DIACYL-GLYCEROL-3-SN-PHOSPHATE / PHOSPHATIDIC ACID / Phosphatidic acid


Mass: 704.998 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C39H77O8P / Feature type: SUBJECT OF INVESTIGATION
#3: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY

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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: Homo-dimer of SLC9C1 transporter / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.294733 MDa / Experimental value: NO
Source (natural)Organism: Strongylocentrotus purpuratus (purple sea urchin)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293F
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE / Humidity: 100 %

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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 FIELDBright-field microscopy / Nominal defocus max: 2000 nm / Nominal defocus min: 600 nm
Image recordingElectron dose: 58.45 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.20.1_4487: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.08 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 1737146 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00316244
ELECTRON MICROSCOPYf_angle_d0.42622054
ELECTRON MICROSCOPYf_dihedral_angle_d6.4912368
ELECTRON MICROSCOPYf_chiral_restr0.0352658
ELECTRON MICROSCOPYf_plane_restr0.0042754

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