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- EMDB-44742: Cryo-EM structure of human Spns1 in complex with LPC (18:1) -

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Basic information

Entry
Database: EMDB / ID: EMD-44742
TitleCryo-EM structure of human Spns1 in complex with LPC (18:1)
Map data
Sample
  • Organelle or cellular component: human Spns1 purified in the presence of extra LPC addition
    • Protein or peptide: mVenus,Maltose/maltodextrin-binding periplasmic protein,Protein spinster homolog 1,Designed ankyrin repeat protein (DARPin)
  • Ligand: (4R,7R,18Z)-4,7-dihydroxy-N,N,N-trimethyl-10-oxo-3,5,9-trioxa-4-phosphaheptacos-18-en-1-aminium 4-oxide
Keywordslysosome / major facilitator superfamily / lysophospholipid transporter / lysophosphatidylcholine / MEMBRANE PROTEIN
Function / homology
Function and homology information


lysophospholipid transport / regulation of lysosomal lumen pH / phospholipid efflux / detection of maltose stimulus / maltose transport complex / carbohydrate transport / transmembrane transporter activity / carbohydrate transmembrane transporter activity / maltose binding / maltose transport ...lysophospholipid transport / regulation of lysosomal lumen pH / phospholipid efflux / detection of maltose stimulus / maltose transport complex / carbohydrate transport / transmembrane transporter activity / carbohydrate transmembrane transporter activity / maltose binding / maltose transport / maltodextrin transmembrane transport / ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing / ATP-binding cassette (ABC) transporter complex / bioluminescence / generation of precursor metabolites and energy / cell chemotaxis / outer membrane-bounded periplasmic space / periplasmic space / mitochondrial inner membrane / lysosomal membrane / DNA damage response / membrane
Similarity search - Function
Protein spinster-like / Major facilitator superfamily / Major Facilitator Superfamily / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / Maltose/Cyclodextrin ABC transporter, substrate-binding protein / Solute-binding family 1, conserved site / Bacterial extracellular solute-binding proteins, family 1 signature. / MFS transporter superfamily / Bacterial extracellular solute-binding protein ...Protein spinster-like / Major facilitator superfamily / Major Facilitator Superfamily / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / Maltose/Cyclodextrin ABC transporter, substrate-binding protein / Solute-binding family 1, conserved site / Bacterial extracellular solute-binding proteins, family 1 signature. / MFS transporter superfamily / Bacterial extracellular solute-binding protein / Bacterial extracellular solute-binding protein / Green fluorescent protein, GFP / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein
Similarity search - Domain/homology
Maltose/maltodextrin-binding periplasmic protein / Green fluorescent protein / Protein spinster homolog 1
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.22 Å
AuthorsChen H / Li X
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)P01 HL160487 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01 GM135343 United States
Welch FoundationI-1957 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2025
Title: Molecular basis of Spns1-mediated lysophospholipid transport from the lysosome.
Authors: Hongwen Chen / Hoa T T Ha / Nadia Elghobashi-Meinhardt / Nhung A Le / Philip Schmiege / Long N Nguyen / Xiaochun Li /
Abstract: Spns1 mediates the rate-limiting efflux of lysophospholipids from the lysosome to the cytosol. Deficiency of Spns1 is associated with embryonic senescence, as well as liver and skeletal muscle ...Spns1 mediates the rate-limiting efflux of lysophospholipids from the lysosome to the cytosol. Deficiency of Spns1 is associated with embryonic senescence, as well as liver and skeletal muscle atrophy in animal models. However, the mechanisms by which Spns1 transports lysophospholipid and proton sensing remain unclear. Here, we present a cryogenic electron microscopy structure of human Spns1 in lysophosphatidylcholine (LPC)-bound lumen-facing conformation. Notably, LPC snugly binds within the luminal-open cavity, where the molecular dynamics simulations reveal that LPC presents a propensity to enter between transmembrane-helices (TM) 5 and 8. Structural comparisons and cell-based transport assays uncover several pivotal residues at TM 5/8 that orchestrate the transport cycle, which are unique to Spns1. Furthermore, we identify a five-residue network that is crucial for proton-sensing by Spns1. Transference of these network residues to Spns2, a sphingosine-1-phosphate uniporter, causes the chimeric Spns2 to be low pH dependent. Our results reveal molecular insights into lysosomal LPC transport and the proton-sensing mechanism by Spns1.
History
DepositionMay 3, 2024-
Header (metadata) releaseJan 1, 2025-
Map releaseJan 1, 2025-
UpdateJan 22, 2025-
Current statusJan 22, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_44742.png

Downloads & links

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Map

FileDownload / File: emd_44742.map.gz / Format: CCP4 / Size: 134.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.74 Å/pix.
x 328 pix.
= 242.064 Å		0.74 Å/pix.
x 328 pix.
= 242.064 Å		0.74 Å/pix.
x 328 pix.
= 242.064 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.738 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.569
Minimum - Maximum-3.6393788 - 4.454293
Average (Standard dev.)0.0018344894 (±0.05999836)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions328328328
Spacing328328328
CellA=B=C: 242.064 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_44742_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_44742_half_map_2.map
Projections & Slices
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Histogram

Histogram (log scale)

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Sample components

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Entire : human Spns1 purified in the presence of extra LPC addition

EntireName: human Spns1 purified in the presence of extra LPC addition
Components
  • Organelle or cellular component: human Spns1 purified in the presence of extra LPC addition
    • Protein or peptide: mVenus,Maltose/maltodextrin-binding periplasmic protein,Protein spinster homolog 1,Designed ankyrin repeat protein (DARPin)
  • Ligand: (4R,7R,18Z)-4,7-dihydroxy-N,N,N-trimethyl-10-oxo-3,5,9-trioxa-4-phosphaheptacos-18-en-1-aminium 4-oxide

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Supramolecule #1: human Spns1 purified in the presence of extra LPC addition

SupramoleculeName: human Spns1 purified in the presence of extra LPC addition
type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: mVenus,Maltose/maltodextrin-binding periplasmic protein,Protein s...

MacromoleculeName: mVenus,Maltose/maltodextrin-binding periplasmic protein,Protein spinster homolog 1,Designed ankyrin repeat protein (DARPin)
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 139.299703 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MVSKGEELFT GVVPILVELD GDVNGHKFSV SGEGEGDATY GKLTLKLICT TGKLPVPWPT LVTTLGYGLQ CFARYPDHMK QHDFFKSAM PEGYVQERTI FFKDDGNYKT RAEVKFEGDT LVNRIELKGI DFKEDGNILG HKLEYNYNSH NVYITADKQK N GIKANFKI ...String:
MVSKGEELFT GVVPILVELD GDVNGHKFSV SGEGEGDATY GKLTLKLICT TGKLPVPWPT LVTTLGYGLQ CFARYPDHMK QHDFFKSAM PEGYVQERTI FFKDDGNYKT RAEVKFEGDT LVNRIELKGI DFKEDGNILG HKLEYNYNSH NVYITADKQK N GIKANFKI RHNIEDGGVQ LADHYQQNTP IGDGPVLLPD NHYLSYQSKL SKDPNEKRDH MVLLEFVTAA GITLGMDELY KL EVLFQGP EEGKLVIWIN GDKGYNGLAE VGKKFEKDTG IKVTVEHPDK LEEKFPQVAA TGDGPDIIFW AHDRFGGYAQ SGL LAEITP DKAFQDKLYP FTWDAVRYNG KLIAYPIAVE ALSLIYNKDL LPNPPKTWEE IPALDKELKA KGKSALMFNL QEPY FTWPL IAADGGYAFK YENGKYDIKD VGVDNAGAKA GLTFLIDLIK NKHMNADTDY SIAEAAFNKG ETAMTINGPW AWSNI DTSK VNYGVTVLPT FKGQPSKPFV GVLSAGINAA SPNKELAKEF LENYLLTDEG LEAVNKDKPL GAVALKSYEE ELVKDP RVA ATMENAQKGE IMPNIPQMSA FWYAVRTAVI NAASGRQTVD AALAAAQAEE EKRKAEEEKR KGRSALIVAV LCYINLL NY MDRFTVAGVL PDIEQFFNIG DSSSGLIQTV FISSYMVLAP VFGYLGDRYN RKYLMCGGIA FWSLVTLGSS FIPGEHFW L LLLTRGLVGV GEASYSTIAP TLIADLFVAD QRSRMLSIFY FAIPVGSGLG YIAGSKVKDM AGDWHWALRV TPGLGVVAV LLLFLVVREP PRGAVERHSD LPPLNPTSWW ADLRALARNP SFVLSSLGFT AVAFVTGSLA LWAPAFLLRS RVVLGETPPC LPGDSCSSS DSLIFGLITC LTGVLGVGLG VEISRRLRHS NPRADPLVCA TGLLGSAPFL FLSLACARGS IVATYIFIFI G ETLLSMNW AIVADILLYV VIPTRRSTAE AFQIVLSHLL GDAGSPYLIG LISDRLRRNW PPSFLSEFRA LQFSLMLCAF VG ALGGAAF LGTAIFIEAD RRRAQLHVQG LLHEAGSTDD RIVVPQRGRS TRVPVASVLI GGGGSGGGGS DLGRKLLEAA RAG QLDEVR ILLANGADVN AADNTGTTPL HLAAYSGHLE IVEVLLKHGA DVDASDVFGY TPLHLAAYWG HLEIVEVLLK NGAD VNAMD SDGMTPLHLA AKWGYLEIVE VLLKHGADVN AQDKFGKTAF DISIDNGNED LAEILQKLNS GRWSHPQFEK

UniProtKB: Green fluorescent protein, Maltose/maltodextrin-binding periplasmic protein, Protein spinster homolog 1

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Macromolecule #2: (4R,7R,18Z)-4,7-dihydroxy-N,N,N-trimethyl-10-oxo-3,5,9-trioxa-4-p...

MacromoleculeName: (4R,7R,18Z)-4,7-dihydroxy-N,N,N-trimethyl-10-oxo-3,5,9-trioxa-4-phosphaheptacos-18-en-1-aminium 4-oxide
type: ligand / ID: 2 / Number of copies: 1 / Formula: 42H
Molecular weightTheoretical: 522.675 Da
Chemical component information

ChemComp-42H:
(4R,7R,18Z)-4,7-dihydroxy-N,N,N-trimethyl-10-oxo-3,5,9-trioxa-4-phosphaheptacos-18-en-1-aminium 4-oxide

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration15 mg/mL
BufferpH: 5.5
GridModel: Quantifoil R1.2/1.3 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.22 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 614781
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: ANGULAR RECONSTITUTION
FSC plot (resolution estimation)

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