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- EMDB-0568: Cryo-EM 3D map of human ATP-citrate lyase N-terminal segment in c... -

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Basic information

Entry
Database: EMDB / ID: EMD-0568
TitleCryo-EM 3D map of human ATP-citrate lyase N-terminal segment in complex with inhibitor NDI-091143
Map dataem-volume_P1
Sample
  • Complex: protein-inhibitor complex
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.11 Å
AuthorsWei J / Tong L
CitationJournal: Nature / Year: 2019
Title: An allosteric mechanism for potent inhibition of human ATP-citrate lyase.
Authors: Jia Wei / Silvana Leit / Jun Kuai / Eric Therrien / Salma Rafi / H James Harwood / Byron DeLaBarre / Liang Tong /
Abstract: ATP-citrate lyase (ACLY) is a central metabolic enzyme and catalyses the ATP-dependent conversion of citrate and coenzyme A (CoA) to oxaloacetate and acetyl-CoA. The acetyl-CoA product is crucial for ...ATP-citrate lyase (ACLY) is a central metabolic enzyme and catalyses the ATP-dependent conversion of citrate and coenzyme A (CoA) to oxaloacetate and acetyl-CoA. The acetyl-CoA product is crucial for the metabolism of fatty acids, the biosynthesis of cholesterol, and the acetylation and prenylation of proteins. There has been considerable interest in ACLY as a target for anti-cancer drugs, because many cancer cells depend on its activity for proliferation. ACLY is also a target against dyslipidaemia and hepatic steatosis, with a compound currently in phase 3 clinical trials. Many inhibitors of ACLY have been reported, but most of them have weak activity. Here we report the development of a series of low nanomolar, small-molecule inhibitors of human ACLY. We have also determined the structure of the full-length human ACLY homo-tetramer in complex with one of these inhibitors (NDI-091143) by cryo-electron microscopy, which reveals an unexpected mechanism of inhibition. The compound is located in an allosteric, mostly hydrophobic cavity next to the citrate-binding site, and requires extensive conformational changes in the enzyme that indirectly disrupt citrate binding. The observed binding mode is supported by and explains the structure-activity relationships of these compounds. This allosteric site greatly enhances the 'druggability' of ACLY and represents an attractive target for the development of new ACLY inhibitors.
History
DepositionFeb 16, 2019-
Header (metadata) releaseMar 13, 2019-
Map releaseMay 1, 2019-
UpdateMay 8, 2019-
Current statusMay 8, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.027
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.027
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_0568.png

Downloads & links

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Map

FileDownload / File: emd_0568.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationem-volume_P1
Voxel sizeX=Y=Z: 1.0605 Å
Density
Contour LevelBy AUTHOR: 0.027 / Movie #1: 0.027
Minimum - Maximum-0.08003193 - 0.12672888
Average (Standard dev.)0.0001709734 (±0.002984444)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 271.488 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.06051.06051.0605
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z271.488271.488271.488
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0800.1270.000

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Supplemental data

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Mask #1

Fileemd_0568_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: em-half-volume P1

Fileemd_0568_half_map_1.map
Annotationem-half-volume_P1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: em-half-volume P2

Fileemd_0568_half_map_2.map
Annotationem-half-volume_P2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : protein-inhibitor complex

EntireName: protein-inhibitor complex
Components
  • Complex: protein-inhibitor complex

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Supramolecule #1: protein-inhibitor complex

SupramoleculeName: protein-inhibitor complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant strain: BL21 STAR

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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

Concentration4.7 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
20.0 mMTris-HClTris-HCl
300.0 mMNaClSodium chloride
2.0 mMDTTDithiothreitol
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK II

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number real images: 2803 / Average exposure time: 10.0 sec. / Average electron dose: 71.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: -0.002 µm / Nominal defocus min: -0.0008 µm / Nominal magnification: 130000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 250156
CTF correctionSoftware - Name: CTFFIND (ver. 4.1)
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.11 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0-beta-2) / Number images used: 63657
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0-beta-2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0-beta-2)
Final 3D classificationSoftware - Name: RELION (ver. 3.0-beta-2)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: AB INITIO MODEL

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