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- PDB-6o0h: Cryo-EM structure of human ATP-citrate lyase in complex with inhi... -

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Basic information

Entry
Database: PDB / ID: 6o0h
TitleCryo-EM structure of human ATP-citrate lyase in complex with inhibitor NDI-091143
ComponentsATP-citrate synthase
Keywordsligase/ligase inhibitor / ATP-citrate lyase / LIGASE / ligase-ligase inhibitor complex
Function / homology
Function and homology information


ATP citrate synthase / ATP citrate synthase activity / Fatty acyl-CoA biosynthesis / citrate metabolic process / ChREBP activates metabolic gene expression / acetyl-CoA biosynthetic process / oxaloacetate metabolic process / coenzyme A metabolic process / lipid biosynthetic process / cholesterol biosynthetic process ...ATP citrate synthase / ATP citrate synthase activity / Fatty acyl-CoA biosynthesis / citrate metabolic process / ChREBP activates metabolic gene expression / acetyl-CoA biosynthetic process / oxaloacetate metabolic process / coenzyme A metabolic process / lipid biosynthetic process / cholesterol biosynthetic process / tricarboxylic acid cycle / fatty acid biosynthetic process / azurophil granule lumen / ficolin-1-rich granule lumen / Neutrophil degranulation / extracellular exosome / extracellular region / nucleoplasm / ATP binding / membrane / metal ion binding / cytosol
Similarity search - Function
ATP-citrate synthase / ATP-citrate synthase, citrate-binding domain / ATP citrate lyase citrate-binding / ATP-citrate lyase/succinyl-CoA ligase, active site / ATP-citrate lyase/succinyl-CoA ligase, conserved site / ATP-citrate lyase / succinyl-CoA ligases family active site. / ATP-citrate lyase / succinyl-CoA ligases family signature 1. / Succinyl-CoA synthetase, beta subunit, conserved site / ATP-citrate lyase / succinyl-CoA ligases family signature 3. / ATP-citrate lyase/succinyl-CoA ligase ...ATP-citrate synthase / ATP-citrate synthase, citrate-binding domain / ATP citrate lyase citrate-binding / ATP-citrate lyase/succinyl-CoA ligase, active site / ATP-citrate lyase/succinyl-CoA ligase, conserved site / ATP-citrate lyase / succinyl-CoA ligases family active site. / ATP-citrate lyase / succinyl-CoA ligases family signature 1. / Succinyl-CoA synthetase, beta subunit, conserved site / ATP-citrate lyase / succinyl-CoA ligases family signature 3. / ATP-citrate lyase/succinyl-CoA ligase / CoA-ligase / CoA binding domain / Succinyl-CoA synthetase-like / CoA binding domain / CoA-binding / Citrate synthase / Citrate synthase-like, small alpha subdomain / Citrate synthase superfamily / Citrate synthase, C-terminal domain / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / Chem-LBG / ATP-citrate synthase
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.67 Å
AuthorsWei, J. / Tong, L.
Funding support United States, 4items
OrganizationGrant numberCountry
Other privateNimbus Therapeutics United States
Other privateSimons Foundation (349247) United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103310 United States
Other governmentNYSTAR United States
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, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 17, 2019Provider: repository / Type: Initial release
Revision 1.1May 8, 2019Group: Data collection / Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.2Jan 8, 2020Group: Author supporting evidence / Data collection / Category: em_imaging / pdbx_audit_support
Item: _em_imaging.nominal_defocus_max / _em_imaging.nominal_defocus_min / _pdbx_audit_support.funding_organization
Revision 1.3Mar 20, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Assembly

Deposited unit
A: ATP-citrate synthase
B: ATP-citrate synthase
C: ATP-citrate synthase
D: ATP-citrate synthase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)494,67312
Polymers491,1484
Non-polymers3,5248
Water0
1


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

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Components

#1: Protein
ATP-citrate synthase / ATP-citrate (pro-S-)-lyase / ACL / Citrate cleavage enzyme


Mass: 122787.102 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ACLY / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 STAR / References: UniProt: P53396, ATP citrate synthase
#2: Chemical
ChemComp-LBG / methyl 3-chloro-5-[(4,6-difluoro[1,1'-biphenyl]-3-yl)sulfamoyl]-4-hydroxybenzoate


Mass: 453.844 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C20H14ClF2NO5S
#3: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE / Adenosine diphosphate


Mass: 427.201 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Comment: ADP, energy-carrying molecule*YM

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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: protein-inhibitor complex / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Escherichia coli (E. coli) / Strain: BL21 STAR
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMTris-HClTrisTris-HClTris1
2300 mMSodium chlorideNaClSodium chloride1
32 mMDithiothreitolDTT1
SpecimenConc.: 4.7 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: C-flat-1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK II / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K

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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 magnification: 130000 X / Nominal defocus max: -2000 nm / Nominal defocus min: -800 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 10 sec. / Electron dose: 71 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 2803

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Processing

EM software
IDNameVersionCategory
1Gautomatch0.56particle selection
2Leginon3.3image acquisition
4CTFFIND4.1CTF correction
7Cootmodel fitting
9RELION3.0-beta-2initial Euler assignment
10RELION3.0-beta-2final Euler assignment
11RELION3.0-beta-2classification
12RELION3.0-beta-23D reconstruction
13PHENIXmodel refinement
CTF correctionType: NONE
Particle selectionNum. of particles selected: 250156
SymmetryPoint symmetry: D2 (2x2 fold dihedral)
3D reconstructionResolution: 3.67 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 11452 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL

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