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- PDB-8g8w: Molecular mechanism of nucleotide inhibition of human uncoupling ... -

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Basic information

Entry
Database: PDB / ID: 8g8w
TitleMolecular mechanism of nucleotide inhibition of human uncoupling protein 1
Components
  • Mitochondrial brown fat uncoupling protein 1
  • Pro-Macrobody 65, Maltose/maltodextrin-binding periplasmic protein chimera
  • Pro-macrobody 71, Maltose/maltodextrin-binding periplasmic protein chimera
KeywordsMEMBRANE PROTEIN / SLC25 / mitochondrial carrier / uncoupling
Function / homology
Function and homology information


purine ribonucleotide binding / cellular response to dehydroepiandrosterone / Mitochondrial Uncoupling / The fatty acid cycling model / oxidative phosphorylation uncoupler activity / mitochondrial transmembrane transport / adaptive thermogenesis / cardiolipin binding / cellular response to fatty acid / regulation of reactive oxygen species biosynthetic process ...purine ribonucleotide binding / cellular response to dehydroepiandrosterone / Mitochondrial Uncoupling / The fatty acid cycling model / oxidative phosphorylation uncoupler activity / mitochondrial transmembrane transport / adaptive thermogenesis / cardiolipin binding / cellular response to fatty acid / regulation of reactive oxygen species biosynthetic process / long-chain fatty acid binding / response to temperature stimulus / diet induced thermogenesis / cellular response to cold / detection of maltose stimulus / maltose transport complex / proton transmembrane transporter activity / carbohydrate transport / carbohydrate transmembrane transporter activity / maltose binding / transmembrane transporter activity / maltose transport / maltodextrin transmembrane transport / brown fat cell differentiation / Transcriptional regulation of brown and beige adipocyte differentiation by EBF2 / cellular response to hormone stimulus / ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing / proton transmembrane transport / ATP-binding cassette (ABC) transporter complex / response to cold / cell chemotaxis / cellular response to reactive oxygen species / response to nutrient levels / GDP binding / positive regulation of cold-induced thermogenesis / outer membrane-bounded periplasmic space / periplasmic space / mitochondrial inner membrane / DNA damage response / regulation of transcription by RNA polymerase II / GTP binding / mitochondrion / membrane
Similarity search - Function
Mitochondrial carrier fold / Mitochondrial carrier domain / : / Mitochondrial carrier protein / Mitochondrial substrate/solute carrier / Mitochondrial carrier domain superfamily / Mitochondrial carrier protein / Solute carrier (Solcar) repeat profile. / Alpha/alpha barrel / Maltose/Cyclodextrin ABC transporter, substrate-binding protein ...Mitochondrial carrier fold / Mitochondrial carrier domain / : / Mitochondrial carrier protein / Mitochondrial substrate/solute carrier / Mitochondrial carrier domain superfamily / Mitochondrial carrier protein / Solute carrier (Solcar) repeat profile. / Alpha/alpha barrel / Maltose/Cyclodextrin ABC transporter, substrate-binding protein / Solute-binding family 1, conserved site / Bacterial extracellular solute-binding proteins, family 1 signature. / Bacterial extracellular solute-binding protein / Bacterial extracellular solute-binding protein / Immunoglobulins / Immunoglobulin-like / Sandwich / Mainly Beta / Mainly Alpha
Similarity search - Domain/homology
CARDIOLIPIN / GUANOSINE-5'-TRIPHOSPHATE / Maltose/maltodextrin-binding periplasmic protein / Mitochondrial brown fat uncoupling protein 1
Similarity search - Component
Biological speciesHomo sapiens (human)
synthetic construct (others)
Escherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsGogoi, P. / Jones, S.A. / Ruprecht, J.J. / King, M.S. / Lee, Y. / Zogg, T. / Pardon, E. / Chand, D. / Steimle, S. / Copeman, D. ...Gogoi, P. / Jones, S.A. / Ruprecht, J.J. / King, M.S. / Lee, Y. / Zogg, T. / Pardon, E. / Chand, D. / Steimle, S. / Copeman, D. / Cotrim, C.A. / Steyaert, J. / Crichton, P.G. / Moiseenkova-Bell, V. / Kunji, E.R.S.
Funding support United States, United Kingdom, European Union, Belgium, 6items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM144120 United States
Medical Research Council (MRC, United Kingdom)MC_UU_00028/2 United Kingdom
Medical Research Council (MRC, United Kingdom)MC_UU_00015/1 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/S00940X/1 United Kingdom
European Union (EU)Instruct-ERIC/ESFRIEuropean Union
Research Foundation - Flanders (FWO) Belgium
CitationJournal: Sci Adv / Year: 2023
Title: Structural basis of purine nucleotide inhibition of human uncoupling protein 1.
Authors: Scott A Jones / Prerana Gogoi / Jonathan J Ruprecht / Martin S King / Yang Lee / Thomas Zögg / Els Pardon / Deepak Chand / Stefan Steimle / Danielle M Copeman / Camila A Cotrim / Jan ...Authors: Scott A Jones / Prerana Gogoi / Jonathan J Ruprecht / Martin S King / Yang Lee / Thomas Zögg / Els Pardon / Deepak Chand / Stefan Steimle / Danielle M Copeman / Camila A Cotrim / Jan Steyaert / Paul G Crichton / Vera Moiseenkova-Bell / Edmund R S Kunji /
Abstract: Mitochondrial uncoupling protein 1 (UCP1) gives brown adipose tissue of mammals its specialized ability to burn calories as heat for thermoregulation. When activated by fatty acids, UCP1 catalyzes ...Mitochondrial uncoupling protein 1 (UCP1) gives brown adipose tissue of mammals its specialized ability to burn calories as heat for thermoregulation. When activated by fatty acids, UCP1 catalyzes the leak of protons across the mitochondrial inner membrane, short-circuiting the mitochondrion to generate heat, bypassing ATP synthesis. In contrast, purine nucleotides bind and inhibit UCP1, regulating proton leak by a molecular mechanism that is unclear. We present the cryo-electron microscopy structure of the GTP-inhibited state of UCP1, which is consistent with its nonconducting state. The purine nucleotide cross-links the transmembrane helices of UCP1 with an extensive interaction network. Our results provide a structural basis for understanding the specificity and pH dependency of the regulatory mechanism. UCP1 has retained all of the key functional and structural features required for a mitochondrial carrier-like transport mechanism. The analysis shows that inhibitor binding prevents the conformational changes that UCP1 uses to facilitate proton leak.
History
DepositionFeb 20, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 7, 2023Provider: repository / Type: Initial release
Revision 1.1Nov 6, 2024Group: Data collection / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Mitochondrial brown fat uncoupling protein 1
B: Pro-macrobody 71, Maltose/maltodextrin-binding periplasmic protein chimera
C: Pro-Macrobody 65, Maltose/maltodextrin-binding periplasmic protein chimera
hetero molecules


Theoretical massNumber of molelcules
Total (without water)146,1048
Polymers140,8463
Non-polymers5,2585
Water00
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Antibody , 2 types, 2 molecules BC

#2: Antibody Pro-macrobody 71, Maltose/maltodextrin-binding periplasmic protein chimera / MMBP / Maltodextrin-binding protein / Maltose-binding protein / MBP


Mass: 53453.910 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others), (gene. exp.) Escherichia coli (strain K12) (bacteria)
Gene: malE, b4034, JW3994 / Production host: Escherichia coli (E. coli) / References: UniProt: P0AEX9
#3: Antibody Pro-Macrobody 65, Maltose/maltodextrin-binding periplasmic protein chimera / MMBP / Maltodextrin-binding protein / Maltose-binding protein / MBP


Mass: 54052.820 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others), (gene. exp.) Escherichia coli (strain K12) (bacteria)
Gene: malE, b4034, JW3994 / Production host: Escherichia coli (E. coli) / References: UniProt: P0AEX9

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Protein / Sugars , 2 types, 2 molecules A

#1: Protein Mitochondrial brown fat uncoupling protein 1 / UCP 1 / Solute carrier family 25 member 7 / Thermogenin


Mass: 33339.629 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: UCP1, SLC25A7, UCP / Production host: Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P25874
#4: Polysaccharide alpha-D-glucopyranose-(1-4)-alpha-D-glucopyranose


Type: oligosaccharide / Mass: 342.297 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpa1-4DGlcpa1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1a_1-5]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][a-D-Glcp]{[(4+1)][a-D-Glcp]{}}LINUCSPDB-CARE

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Non-polymers , 2 types, 4 molecules

#5: Chemical ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE


Mass: 523.180 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O14P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: GTP, energy-carrying molecule*YM
#6: Chemical ChemComp-CDL / CARDIOLIPIN / DIPHOSPHATIDYL GLYCEROL / BIS-(1,2-DIACYL-SN-GLYCERO-3-PHOSPHO)-1',3'-SN-GLYCEROL


Mass: 1464.043 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C81H156O17P2 / Comment: phospholipid*YM

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Details

Has ligand of interestY
Has protein modificationY

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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: Complex of human uncoupling protein 1 with two promacrobodies
Type: COMPLEX / Entity ID: #1-#3 / Source: MULTIPLE SOURCES
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Saccharomyces cerevisiae (brewer's yeast)
Buffer solutionpH: 6
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMMESC6H13NO4S1
2150 mMSodium chlorideNaCl1
31 mMTCEPC9H15O6P1
40.02 %DMNGC43H80O221
50.02 mg/mLTetraoleoyl cardiolipinC81H150O17P21
62 mMGTPC10H16N5O14P31
72 mMD-maltoseC12H22O111
SpecimenConc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.14 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 FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 72.9 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 203799 / Symmetry type: POINT

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