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- EMDB-42661: Site-one protease without SPRING -

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Basic information

Entry
Database: EMDB / ID: EMD-42661
TitleSite-one protease without SPRING
Map dataPrimary, unsharpened map for S1P
Sample
  • Complex: Matured site-1 protease
    • Protein or peptide: Membrane-bound transcription factor site-1 protease
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: water
Keywordsserine protease cholesterol metabolism zymogen activation Protein complex glycoprotein secretory pathway / SIGNALING PROTEIN
Function / homology
Function and homology information


site-1 protease / CREB3 factors activate genes / ATF6-mediated unfolded protein response / regulation of cholesterol biosynthetic process / ATF6 (ATF6-alpha) activates chaperones / ATF6B (ATF6-beta) activates chaperones / Assembly of active LPL and LIPC lipase complexes / membrane protein intracellular domain proteolysis / regulation of vesicle-mediated transport / Regulation of cholesterol biosynthesis by SREBP (SREBF) ...site-1 protease / CREB3 factors activate genes / ATF6-mediated unfolded protein response / regulation of cholesterol biosynthetic process / ATF6 (ATF6-alpha) activates chaperones / ATF6B (ATF6-beta) activates chaperones / Assembly of active LPL and LIPC lipase complexes / membrane protein intracellular domain proteolysis / regulation of vesicle-mediated transport / Regulation of cholesterol biosynthesis by SREBP (SREBF) / Golgi stack / lysosome organization / mitotic G2 DNA damage checkpoint signaling / protein maturation / endoplasmic reticulum unfolded protein response / cholesterol metabolic process / response to endoplasmic reticulum stress / Post-translational protein phosphorylation / protein processing / Regulation of Insulin-like Growth Factor (IGF) transport and uptake by Insulin-like Growth Factor Binding Proteins (IGFBPs) / endoplasmic reticulum lumen / Golgi membrane / serine-type endopeptidase activity / endoplasmic reticulum membrane / Golgi apparatus / proteolysis
Similarity search - Function
Site-1 peptidase catalytic domain / Peptidase S8, subtilisin, His-active site / Serine proteases, subtilase family, histidine active site. / Serine proteases, subtilase family, serine active site. / Peptidase S8, subtilisin, Ser-active site / Serine proteases, subtilase domain profile. / Peptidase S8, subtilisin-related / Peptidase S8/S53 domain superfamily / Subtilase family / Peptidase S8/S53 domain
Similarity search - Domain/homology
Membrane-bound transcription factor site-1 protease
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.27 Å
AuthorsKober DL
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R00GM141261 United States
CitationJournal: Nat Commun / Year: 2024
Title: SPRING licenses S1P-mediated cleavage of SREBP2 by displacing an inhibitory pro-domain.
Authors: Sebastian Hendrix / Vincent Dartigue / Hailee Hall / Shrankhla Bawaria / Jenina Kingma / Bilkish Bajaj / Noam Zelcer / Daniel L Kober /
Abstract: Site-one protease (S1P) conducts the first of two cleavage events in the Golgi to activate Sterol regulatory element binding proteins (SREBPs) and upregulate lipogenic transcription. S1P is also ...Site-one protease (S1P) conducts the first of two cleavage events in the Golgi to activate Sterol regulatory element binding proteins (SREBPs) and upregulate lipogenic transcription. S1P is also required for a wide array of additional signaling pathways. A zymogen serine protease, S1P matures through autoproteolysis of two pro-domains, with one cleavage event in the endoplasmic reticulum (ER) and the other in the Golgi. We recently identified the SREBP regulating gene, (SPRING), which enhances S1P maturation and is necessary for SREBP signaling. Here, we report the cryo-EM structures of S1P and S1P-SPRING at sub-2.5 Å resolution. SPRING activates S1P by dislodging its inhibitory pro-domain and stabilizing intra-domain contacts. Functionally, SPRING licenses S1P to cleave its cognate substrate, SREBP2. Our findings reveal an activation mechanism for S1P and provide insights into how spatial control of S1P activity underpins cholesterol homeostasis.
History
DepositionNov 6, 2023-
Header (metadata) releaseJul 10, 2024-
Map releaseJul 10, 2024-
UpdateJul 24, 2024-
Current statusJul 24, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_42661.png

Downloads & links

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Map

FileDownload / File: emd_42661.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationPrimary, unsharpened map for S1P
Voxel sizeX=Y=Z: 0.738 Å
Density
Contour LevelBy AUTHOR: 0.25
Minimum - Maximum-0.96416175 - 1.3770901
Average (Standard dev.)-0.00014512613 (±0.01758762)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 295.19998 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: sharpened map (B factor = -62)

Fileemd_42661_additional_1.map
Annotationsharpened map (B factor = -62)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map A

Fileemd_42661_half_map_1.map
Annotationhalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map B

Fileemd_42661_half_map_2.map
Annotationhalf map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Matured site-1 protease

EntireName: Matured site-1 protease
Components
  • Complex: Matured site-1 protease
    • Protein or peptide: Membrane-bound transcription factor site-1 protease
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: water

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Supramolecule #1: Matured site-1 protease

SupramoleculeName: Matured site-1 protease / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: FLAG-S1P-His purified using NiNTA chromatography and gel filtration.
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 120 KDa

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Macromolecule #1: Membrane-bound transcription factor site-1 protease

MacromoleculeName: Membrane-bound transcription factor site-1 protease / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: site-1 protease
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 113.883789 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: METDTLLLWV LLLWVPGSTG DYKDDDDKDR LEKKSFEKAP CPGCSHLTLK VEFSSTVVEY EYIVAFNGYF TAKARNSFIS SALKSSEVD NWRIIPRNNP SSDYPSDFEV IQIKEKQKAG LLTLEDHPNI KRVTPQRKVF RSLKYAESDP TVPCNETRWS Q KWQSSRPL ...String:
METDTLLLWV LLLWVPGSTG DYKDDDDKDR LEKKSFEKAP CPGCSHLTLK VEFSSTVVEY EYIVAFNGYF TAKARNSFIS SALKSSEVD NWRIIPRNNP SSDYPSDFEV IQIKEKQKAG LLTLEDHPNI KRVTPQRKVF RSLKYAESDP TVPCNETRWS Q KWQSSRPL RRASLSLGSG FWHATGRHSS RRLLRAIPRQ VAQTLQADVL WQMGYTGANV RVAVFDTGLS EKHPHFKNVK ER TNWTNER TLDDGLGHGT FVAGVIASMR ECQGFAPDAE LHIFRVFTNN QVSYTSWFLD AFNYAILKKI DVLNLSIGGP DFM DHPFVD KVWELTANNV IMVSAIGNDG PLYGTLNNPA DQMDVIGVGG IDFEDNIARF SSRGMTTWEL PGGYGRMKPD IVTY GAGVR GSGVKGGCRA LSGTSVASPV VAGAVTLLVS TVQKRELVNP ASMKQALIAS ARRLPGVNMF EQGHGKLDLL RAYQI LNSY KPQASLSPSY IDLTECPYMW PYCSQPIYYG GMPTVVNVTI LNGMGVTGRI VDKPDWQPYL PQNGDNIEVA FSYSSV LWP WSGYLAISIS VTKKAASWEG IAQGHVMITV ASPAETESKN GAEQTSTVKL PIKVKIIPTP PRSKRVLWDQ YHNLRYP PG YFPRDNLRMK NDPLDWNGDH IHTNFRDMYQ HLRSMGYFVE VLGAPFTCFD ASQYGTLLMV DSEEEYFPEE IAKLRRDV D NGLSLVIFSD WYNTSVMRKV KFYDENTRQW WMPDTGGANI PALNELLSVW NMGFSDGLYE GEFTLANHDM YYASGCSIA KFPEDGVVIT QTFKDQGLEV LKQETAVVEN VPILGLYQIP AEGGGRIVLY GDSNCLDDSH RQKDCFWLLD ALLQYTSYGV TPPSLSHSG NRQRPPSGAG SVTPERMEGN HLHRYSKVLE AHLGDPKPRP LPACPRLSWA KPQPLNETAP SNLWKHQKLL S IDLDKVVL PNFRSNRPQV RPLSPGESGA WDIPGGIMPG RYNQEVHHHH HHHHHH

UniProtKB: Membrane-bound transcription factor site-1 protease

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Macromolecule #2: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 2 / Number of copies: 1 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #3: water

MacromoleculeName: water / type: ligand / ID: 3 / Number of copies: 97 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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

Concentration0.6 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
150.0 mMNaClsodium chloride
50.0 mMHEPES-NaOHHEPES buffered with sodium hydroxide

Details: 50 mM HEPES-NaOH and 150 mM NaCl
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris / Energy filter - Slit width: 5 eV
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Number grids imaged: 1 / Number real images: 6139 / Average exposure time: 3.5 sec. / Average electron dose: 55.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.6 µm / Nominal defocus min: 0.6 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: 2.27 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2) / Number images used: 659523
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2)
FSC plot (resolution estimation)

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