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- EMDB-17691: 60-meric complex of dihydrolipoamide acetyltransferase (E2) of th... -

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Basic information

Entry
Database: EMDB / ID: EMD-17691
Title60-meric complex of dihydrolipoamide acetyltransferase (E2) of the human pyruvate dehydrogenase complex (icosahedral symmetry)
Map data
Sample
  • Complex: 60-meric complex of dihydrolipoamide acetyltransferase (E2) of the human pyruvate dehydrogenase complex
    • Protein or peptide: Dihydrolipoyllysine-residue acetyltransferase component of pyruvate dehydrogenase complex, mitochondrial
Keywordspyruvate dehydrogenase complex / PDHc / E2 / cryo-EM / TRANSFERASE
Function / homology
Function and homology information


PDH complex synthesizes acetyl-CoA from PYR / dihydrolipoyllysine-residue acetyltransferase / dihydrolipoyllysine-residue acetyltransferase activity / Regulation of pyruvate dehydrogenase (PDH) complex / pyruvate catabolic process / Protein lipoylation / pyruvate decarboxylation to acetyl-CoA / pyruvate dehydrogenase complex / Signaling by Retinoic Acid / tricarboxylic acid cycle ...PDH complex synthesizes acetyl-CoA from PYR / dihydrolipoyllysine-residue acetyltransferase / dihydrolipoyllysine-residue acetyltransferase activity / Regulation of pyruvate dehydrogenase (PDH) complex / pyruvate catabolic process / Protein lipoylation / pyruvate decarboxylation to acetyl-CoA / pyruvate dehydrogenase complex / Signaling by Retinoic Acid / tricarboxylic acid cycle / glucose metabolic process / mitochondrial matrix / mitochondrion / identical protein binding
Similarity search - Function
Dihydrolipoyllysine-residue acetyltransferase component of pyruvate dehydrogenase complex / Dihydrolipoamide acetyltransferase/Pyruvate dehydrogenase protein X component / Peripheral subunit-binding domain / e3 binding domain / Peripheral subunit-binding (PSBD) domain profile. / E3-binding domain superfamily / 2-oxo acid dehydrogenase, lipoyl-binding site / 2-oxo acid dehydrogenases acyltransferase component lipoyl binding site. / 2-oxoacid dehydrogenase acyltransferase, catalytic domain / 2-oxoacid dehydrogenases acyltransferase (catalytic domain) ...Dihydrolipoyllysine-residue acetyltransferase component of pyruvate dehydrogenase complex / Dihydrolipoamide acetyltransferase/Pyruvate dehydrogenase protein X component / Peripheral subunit-binding domain / e3 binding domain / Peripheral subunit-binding (PSBD) domain profile. / E3-binding domain superfamily / 2-oxo acid dehydrogenase, lipoyl-binding site / 2-oxo acid dehydrogenases acyltransferase component lipoyl binding site. / 2-oxoacid dehydrogenase acyltransferase, catalytic domain / 2-oxoacid dehydrogenases acyltransferase (catalytic domain) / Biotin-requiring enzyme / Biotinyl/lipoyl domain profile. / Biotin/lipoyl attachment / Single hybrid motif / Chloramphenicol acetyltransferase-like domain superfamily
Similarity search - Domain/homology
Dihydrolipoyllysine-residue acetyltransferase component of pyruvate dehydrogenase complex, mitochondrial
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsZdanowicz R / Afanasyev P / Boehringer D / Glockshuber R
Funding support Switzerland, 1 items
OrganizationGrant numberCountry
Swiss National Science Foundation Switzerland
CitationJournal: Sci Adv / Year: 2024
Title: Stoichiometry and architecture of the human pyruvate dehydrogenase complex.
Authors: Rafal Zdanowicz / Pavel Afanasyev / Adam Pruška / Julian A Harrison / Christoph Giese / Daniel Boehringer / Alexander Leitner / Renato Zenobi / Rudi Glockshuber /
Abstract: The pyruvate dehydrogenase complex (PDHc) is a key megaenzyme linking glycolysis with the citric acid cycle. In mammalian PDHc, dihydrolipoamide acetyltransferase (E2) and the dihydrolipoamide ...The pyruvate dehydrogenase complex (PDHc) is a key megaenzyme linking glycolysis with the citric acid cycle. In mammalian PDHc, dihydrolipoamide acetyltransferase (E2) and the dihydrolipoamide dehydrogenase-binding protein (E3BP) form a 60-subunit core that associates with the peripheral subunits pyruvate dehydrogenase (E1) and dihydrolipoamide dehydrogenase (E3). The structure and stoichiometry of the fully assembled, mammalian PDHc or its core remained elusive. Here, we demonstrate that the human PDHc core is formed by 48 E2 copies that bind 48 E1 heterotetramers and 12 E3BP copies that bind 12 E3 homodimers. Cryo-electron microscopy, together with native and cross-linking mass spectrometry, confirmed a core model in which 8 E2 homotrimers and 12 E2-E2-E3BP heterotrimers assemble into a pseudoicosahedral particle such that the 12 E3BP molecules form six E3BP-E3BP intertrimer interfaces distributed tetrahedrally within the 60-subunit core. The even distribution of E3 subunits in the peripheral shell of PDHc guarantees maximum enzymatic activity of the megaenzyme.
History
DepositionJun 22, 2023-
Header (metadata) releaseJul 10, 2024-
Map releaseJul 10, 2024-
UpdateJul 31, 2024-
Current statusJul 31, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_17691.png

Downloads & links

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Map

FileDownload / File: emd_17691.map.gz / Format: CCP4 / Size: 371.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 460 pix.
= 386.4 Å		0.84 Å/pix.
x 460 pix.
= 386.4 Å		0.84 Å/pix.
x 460 pix.
= 386.4 Å

Surface

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Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.84 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-2.6417093 - 4.432757
Average (Standard dev.)0.02035436 (±0.14072537)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions460460460
Spacing460460460
CellA=B=C: 386.4 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_17691_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_17691_half_map_2.map
Projections & Slices
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Sample components

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Entire : 60-meric complex of dihydrolipoamide acetyltransferase (E2) of th...

EntireName: 60-meric complex of dihydrolipoamide acetyltransferase (E2) of the human pyruvate dehydrogenase complex
Components
  • Complex: 60-meric complex of dihydrolipoamide acetyltransferase (E2) of the human pyruvate dehydrogenase complex
    • Protein or peptide: Dihydrolipoyllysine-residue acetyltransferase component of pyruvate dehydrogenase complex, mitochondrial

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Supramolecule #1: 60-meric complex of dihydrolipoamide acetyltransferase (E2) of th...

SupramoleculeName: 60-meric complex of dihydrolipoamide acetyltransferase (E2) of the human pyruvate dehydrogenase complex
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 3.6 MDa

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Macromolecule #1: Dihydrolipoyllysine-residue acetyltransferase component of pyruva...

MacromoleculeName: Dihydrolipoyllysine-residue acetyltransferase component of pyruvate dehydrogenase complex, mitochondrial
type: protein_or_peptide / ID: 1 / Number of copies: 60 / Enantiomer: LEVO / EC number: dihydrolipoyllysine-residue acetyltransferase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 59.811832 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MSLPPHQKVP LPSLSPTMQA GTIARWEKKE GDKINEGDLI AEVETDKATV GFESLEECYM AKILVAEGTR DVPIGAIICI TVGKPEDIE AFKNYTLDSS AAPTPQAAPA PTPAATASPP TPSAQAPGSS YPPHMQVLLP ALSPTMTMGT VQRWEKKVGE K LSEGDLLA ...String:
MSLPPHQKVP LPSLSPTMQA GTIARWEKKE GDKINEGDLI AEVETDKATV GFESLEECYM AKILVAEGTR DVPIGAIICI TVGKPEDIE AFKNYTLDSS AAPTPQAAPA PTPAATASPP TPSAQAPGSS YPPHMQVLLP ALSPTMTMGT VQRWEKKVGE K LSEGDLLA EIETDKATIG FEVQEEGYLA KILVPEGTRD VPLGTPLCII VEKEADISAF ADYRPTEVTD LKPQVPPPTP PP VAAVPPT PQPLAPTPSA PCPATPAGPK GRVFVSPLAK KLAVEKGIDL TQVKGTGPDG RITKKDIDSF VPSKVAPAPA AVV PPTGPG MAPVPTGVFT DIPISNIRRV IAQRLMQSKQ TIPHYYLSID VNMGEVLLVR KELNKILEGR SKISVNDFII KASA LACLK VPEANSSWMD TVIRQNHVVD VSVAVSTPAG LITPIVFNAH IKGVETIAND VVSLATKARE GKLQPHEFQG GTFTI SNLG MFGIKNFSAI INPPQACILA IGASEDKLVP ADNEKGFDVA SMMSVTLSCD HRVVDGAVGA QWLAEFRKYL EKPITM LL

UniProtKB: Dihydrolipoyllysine-residue acetyltransferase component of pyruvate dehydrogenase complex, mitochondrial

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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

BufferpH: 7.4
GridModel: Quantifoil R2/2 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 80.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.8000000000000003 µm / Nominal defocus min: 1.6 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER / Details: cryoSPARC Ab-Initio model
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.2) / Number images used: 83959
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.2)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6ct0


Chain - Source name: PDB / Chain - Initial model type: experimental model
Output model

PDB-8piu:
60-meric complex of dihydrolipoamide acetyltransferase (E2) of the human pyruvate dehydrogenase complex

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