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- EMDB-28818: Structure of yeast F1-ATPase determined with 100 micromolar cruen... -

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Basic information

Entry
Database: EMDB / ID: EMD-28818
TitleStructure of yeast F1-ATPase determined with 100 micromolar cruentaren A
Map dataSharpened map.
Sample
  • Complex: Yeast F1-ATPase with 100 micromolar cruentaren A
    • Protein or peptide: ATP synthase subunit alpha
    • Protein or peptide: ATP synthase subunit beta
    • Protein or peptide: ATP synthase subunit gamma
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: Cruentaren A
KeywordsF1-ATPase / ATP Synthase / cruentaren A / drug development / HYDROLASE
Function / homology
Function and homology information


proton-transporting ATP synthase complex, catalytic core F(1) / H+-transporting two-sector ATPase / proton-transporting ATPase activity, rotational mechanism / proton-transporting ATP synthase activity, rotational mechanism / mitochondrial inner membrane / ATP hydrolysis activity / ATP binding
Similarity search - Function
ATP synthase, F1 complex, gamma subunit conserved site / ATP synthase gamma subunit signature. / ATP synthase, F1 complex, beta subunit / ATP synthase, alpha subunit, C-terminal domain superfamily / ATP synthase, F1 complex, gamma subunit / ATP synthase, F1 complex, gamma subunit superfamily / ATP synthase / ATP synthase, alpha subunit, C-terminal / ATP synthase, F1 complex, alpha subunit / ATP synthase, F1 complex, alpha subunit nucleotide-binding domain ...ATP synthase, F1 complex, gamma subunit conserved site / ATP synthase gamma subunit signature. / ATP synthase, F1 complex, beta subunit / ATP synthase, alpha subunit, C-terminal domain superfamily / ATP synthase, F1 complex, gamma subunit / ATP synthase, F1 complex, gamma subunit superfamily / ATP synthase / ATP synthase, alpha subunit, C-terminal / ATP synthase, F1 complex, alpha subunit / ATP synthase, F1 complex, alpha subunit nucleotide-binding domain / ATP synthase alpha/beta chain, C terminal domain / ATPase, F1/V1 complex, beta/alpha subunit, C-terminal / ATP synthase subunit alpha, N-terminal domain-like superfamily / ATPase, F1/V1/A1 complex, alpha/beta subunit, N-terminal domain superfamily / ATPase, F1/V1/A1 complex, alpha/beta subunit, N-terminal domain / ATP synthase alpha/beta family, beta-barrel domain / ATPase, alpha/beta subunit, nucleotide-binding domain, active site / ATP synthase alpha and beta subunits signature. / ATPase, F1/V1/A1 complex, alpha/beta subunit, nucleotide-binding domain / ATP synthase alpha/beta family, nucleotide-binding domain / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ATP synthase subunit beta / ATP synthase subunit gamma / ATP synthase subunit alpha
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsGuo H / Rubinstein JL
Funding support Canada, United States, 5 items
OrganizationGrant numberCountry
Canada Research ChairsElectron Cryomicroscopy Canada
Canadian Institutes of Health Research (CIHR)PJT162186 Canada
Canada Foundation for Innovation Canada
National Institutes of Health/National Cancer Institute (NIH/NCI)R01 CA216919 United States
National Institutes of Health/National Center for Advancing Translational Sciences (NIH/NCATS)UL1TR002529 United States
CitationJournal: Chemistry / Year: 2023
Title: CryoEM Structure with ATP Synthase Enables Late-Stage Diversification of Cruentaren A.
Authors: Xiaozheng Dou / Hui Guo / Terin D'Amico / Leah Abdallah / Chitra Subramanian / Bhargav A Patel / Mark Cohen / John L Rubinstein / Brian S J Blagg /
Abstract: Cruentaren A is a natural product that exhibits potent antiproliferative activity against various cancer cell lines, yet its binding site within ATP synthase remained unknown, thus limiting the ...Cruentaren A is a natural product that exhibits potent antiproliferative activity against various cancer cell lines, yet its binding site within ATP synthase remained unknown, thus limiting the development of improved analogues as anticancer agents. Herein, we report the cryogenic electron microscopy (cryoEM) structure of cruentaren A bound to ATP synthase, which allowed the design of new inhibitors through semisynthetic modification. Examples of cruentaren A derivatives include a trans-alkene isomer, which was found to exhibit similar activity to cruentaren A against three cancer cell lines as well as several other analogues that retained potent inhibitory activity. Together, these studies provide a foundation for the generation of cruentaren A derivatives as potential therapeutics for the treatment of cancer.
History
DepositionNov 8, 2022-
Header (metadata) releaseApr 5, 2023-
Map releaseApr 5, 2023-
UpdateApr 24, 2024-
Current statusApr 24, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_28818.png

Downloads & links

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Map

FileDownload / File: emd_28818.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened map.
Voxel sizeX=Y=Z: 1.04622 Å
Density
Contour LevelBy AUTHOR: 1.1
Minimum - Maximum-7.164443 - 10.720364999999999
Average (Standard dev.)0.00116477 (±0.18358803)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 334.79037 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_28818_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Unsharpened map.

Fileemd_28818_additional_1.map
AnnotationUnsharpened map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 1.

Fileemd_28818_half_map_1.map
AnnotationHalf map 1.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 2.

Fileemd_28818_half_map_2.map
AnnotationHalf map 2.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Yeast F1-ATPase with 100 micromolar cruentaren A

EntireName: Yeast F1-ATPase with 100 micromolar cruentaren A
Components
  • Complex: Yeast F1-ATPase with 100 micromolar cruentaren A
    • Protein or peptide: ATP synthase subunit alpha
    • Protein or peptide: ATP synthase subunit beta
    • Protein or peptide: ATP synthase subunit gamma
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: Cruentaren A

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Supramolecule #1: Yeast F1-ATPase with 100 micromolar cruentaren A

SupramoleculeName: Yeast F1-ATPase with 100 micromolar cruentaren A / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: USY006
Molecular weightTheoretical: 400 KDa

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Macromolecule #1: ATP synthase subunit alpha

MacromoleculeName: ATP synthase subunit alpha / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 52.376539 KDa
SequenceString: NLNETGRVLA VGDGIARVFG LNNIQAEELV EFSSGVKGMA LNLEPGQVGI VLFGSDRLVK EGELVKRTGN IVDVPVGPGL LGRVVDALG NPIDGKGPID AAGRSRAQVK APGILPRRSV HEPVQTGLKA VDALVPIGRG QRELIIGDRQ TGKTAVALDT I LNQKRWNN ...String:
NLNETGRVLA VGDGIARVFG LNNIQAEELV EFSSGVKGMA LNLEPGQVGI VLFGSDRLVK EGELVKRTGN IVDVPVGPGL LGRVVDALG NPIDGKGPID AAGRSRAQVK APGILPRRSV HEPVQTGLKA VDALVPIGRG QRELIIGDRQ TGKTAVALDT I LNQKRWNN GSDESKKLYC VYVAVGQKRS TVAQLVQTLE QHDAMKYSII VAATASEAAP LQYLAPFTAA SIGEWFRDNG KH ALIVYDD LSKQAVAYRQ LSLLLRRPPG REAYPGDVFY LHSRLLERAA KLSEKEGSGS LTALPVIETQ GGDVSAYIPT NVI SITDGQ IFLEAELFYK GIRPAINVGL SVSRVGSAAQ VKALKQVAGS LKLFLAQYRE VAAFAQFGSD LDASTKQTLV RGER LTQLL KQNQYSPLAT EEQVPLIYAG VNGHLDGIEL SRIGEFESSF LSYLKSNHNE LLTEIREKGE LSKELLASLK SATES FVAT F

UniProtKB: ATP synthase subunit alpha

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Macromolecule #2: ATP synthase subunit beta

MacromoleculeName: ATP synthase subunit beta / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO / EC number: H+-transporting two-sector ATPase
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 50.379277 KDa
SequenceString: PITGKVTAVI GAIVDVHFEQ SELPAILNAL EIKTPQGKLV LEVAQHLGEN TVRTIAMDGT EGLVRGEKVL DTGGPISVPV GRETLGRII NVIGEPIDER GPIKSKLRKP IHADPPSFAE QSTSAEILET GIKVVDLLAP YARGGKIGLF GGAGVGKTVF I QELINNIA ...String:
PITGKVTAVI GAIVDVHFEQ SELPAILNAL EIKTPQGKLV LEVAQHLGEN TVRTIAMDGT EGLVRGEKVL DTGGPISVPV GRETLGRII NVIGEPIDER GPIKSKLRKP IHADPPSFAE QSTSAEILET GIKVVDLLAP YARGGKIGLF GGAGVGKTVF I QELINNIA KAHGGFSVFT GVGERTREGN DLYREMKETG VINLEGESKV ALVFGQMNEP PGARARVALT GLTIAEYFRD EE GQDVLLF IDNIFRFTQA GSEVSALLGR IPSAVGYQPT LATDMGLLQE RITTTKKGSV TSVQAVYVPA DDLTDPAPAT TFA HLDATT VLSRGISELG IYPAVDPLDS KSRLLDAAVV GQEHYDVASK VQETLQTYKS LQDIIAILGM DELSEQDKLT VERA RKIQR FLSQPFAVAE VFTGIPGKLV RLKDTVASFK AVLEGKYDNI PEHAFYMVGG IEDVVAKAEK LAAE

UniProtKB: ATP synthase subunit beta

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Macromolecule #3: ATP synthase subunit gamma

MacromoleculeName: ATP synthase subunit gamma / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 30.399875 KDa
SequenceString: ATLKEVEMRL KSIKNIEKIT KTMKIVASTR LSKAEKAKIS AKKMDEAEQL FYKNAETKNL DVEATETGAP KELIVAITSD KGLCGSIHS QLAKAVRRHL NDQPNADIVT IGDKIKMQLL RTHPNNIKLS INGIGKDAPT FQESALIADK LLSVMKAGTY P KISIFYND ...String:
ATLKEVEMRL KSIKNIEKIT KTMKIVASTR LSKAEKAKIS AKKMDEAEQL FYKNAETKNL DVEATETGAP KELIVAITSD KGLCGSIHS QLAKAVRRHL NDQPNADIVT IGDKIKMQLL RTHPNNIKLS INGIGKDAPT FQESALIADK LLSVMKAGTY P KISIFYND PVSSLSFEPS EKPIFNAKTI EQSPSFGKFE IDTDANVPRD LFEYTLANQM LTAMAQGYAA EISARRNAMD NA SKNAGDM INRYSILYNR TRQAVITNEL VDIITGAS

UniProtKB: ATP synthase subunit gamma

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Macromolecule #4: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 4 / Number of copies: 5 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM / Adenosine triphosphate

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Macromolecule #5: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 5 / Number of copies: 5 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #6: Cruentaren A

MacromoleculeName: Cruentaren A / type: ligand / ID: 6 / Number of copies: 2 / Formula: XBC
Molecular weightTheoretical: 589.76 Da
Chemical component information

ChemComp-XBC:
Cruentaren A

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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: 7.4
GridModel: Homemade / Material: COPPER/RHODIUM / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Support film - Film thickness: 35 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 120 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: LEICA EM GP

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated magnification: 133815 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 75000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number real images: 5814 / Average exposure time: 8.4 sec. / Average electron dose: 42.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1352159
Startup modelType of model: OTHER
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.1.0)
Final 3D classificationNumber classes: 3 / Avg.num./class: 233814 / Software - Name: cryoSPARC (ver. 3.1.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.1.0)
Final reconstructionNumber classes used: 3 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.1.0) / Number images used: 701444
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

2hld


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: RECIPROCAL / Protocol: RIGID BODY FIT
Output model

PDB-8f2k:
Structure of yeast F1-ATPase determined with 100 micromolar cruentaren A

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