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- EMDB-47956: Cryo-EM model of E. coli aspartate transcarbamoylase in the ligan... -

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Basic information

Entry
Database: EMDB / ID: EMD-47956
TitleCryo-EM model of E. coli aspartate transcarbamoylase in the ligand-free T-state
Map data
Sample
  • Complex: Heterododecameric aspartate transcarbamoylase
    • Protein or peptide: Aspartate carbamoyltransferase regulatory chain
    • Protein or peptide: Aspartate carbamoyltransferase
  • Ligand: ZINC ION
KeywordsComplex / Allostery / ATCase / T-state / CYTOSOLIC PROTEIN
Function / homology
Function and homology information


aspartate carbamoyltransferase complex / pyrimidine nucleotide biosynthetic process / aspartate carbamoyltransferase / aspartate carbamoyltransferase activity / amino acid metabolic process / amino acid binding / 'de novo' UMP biosynthetic process / 'de novo' pyrimidine nucleobase biosynthetic process / transferase activity / metal ion binding / cytosol
Similarity search - Function
Aspartate transcarbamylase regulatory subunit / Aspartate carbamoyltransferase regulatory subunit, C-terminal / Aspartate carbamoyltransferase regulatory subunit, N-terminal / Aspartate carbamoyltransferase regulatory subunit, C-terminal domain superfamily / Aspartate carbamoyltransferase regulatory subunit, N-terminal domain superfamily / Aspartate carbamoyltransferase regulatory chain, allosteric domain / Aspartate carbamoyltransferase regulatory chain, metal binding domain / Aspartate carbamoyltransferase / Aspartate and ornithine carbamoyltransferases signature. / Aspartate/ornithine carbamoyltransferase ...Aspartate transcarbamylase regulatory subunit / Aspartate carbamoyltransferase regulatory subunit, C-terminal / Aspartate carbamoyltransferase regulatory subunit, N-terminal / Aspartate carbamoyltransferase regulatory subunit, C-terminal domain superfamily / Aspartate carbamoyltransferase regulatory subunit, N-terminal domain superfamily / Aspartate carbamoyltransferase regulatory chain, allosteric domain / Aspartate carbamoyltransferase regulatory chain, metal binding domain / Aspartate carbamoyltransferase / Aspartate and ornithine carbamoyltransferases signature. / Aspartate/ornithine carbamoyltransferase / Aspartate/ornithine carbamoyltransferase, Asp/Orn-binding domain / Aspartate/ornithine carbamoyltransferase, carbamoyl-P binding / Aspartate/ornithine carbamoyltransferase superfamily / Aspartate/ornithine carbamoyltransferase, Asp/Orn binding domain / Aspartate/ornithine carbamoyltransferase, carbamoyl-P binding domain
Similarity search - Domain/homology
Aspartate carbamoyltransferase / Aspartate carbamoyltransferase regulatory chain
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.62 Å
AuthorsMiller RC / Ando N
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM124847 United States
National Science Foundation (NSF, United States)DMR-1719875 United States
CitationJournal: Nat Commun / Year: 2026
Title: Cooperativity in E. coli aspartate transcarbamoylase is tuned by allosteric breathing.
Authors: Robert C Miller / Michael G Patterson / Neti Bhatt / Xiaokun Pei / Nozomi Ando /
Abstract: Aspartate transcarbamoylase (ATCase) from Escherichia coli catalyzes a key step in pyrimidine nucleotide biosynthesis and has long served as a model for allosteric regulation. Despite decades of ...Aspartate transcarbamoylase (ATCase) from Escherichia coli catalyzes a key step in pyrimidine nucleotide biosynthesis and has long served as a model for allosteric regulation. Despite decades of study, how nucleotide binding at distant regulatory sites controls cooperativity between active sites remained unresolved. Here we show that ATCase does not simply interconvert between two conformations, as traditionally depicted, but instead samples a continuum of conformations that tune enzyme cooperativity. Using complementary cryo-electron microscopy, small-angle X-ray scattering, and crystallography under conditions that ensure full assembly of the allosteric sites, we show that ATCase behaves like a flexible balloon whose global "breathing" motions directly regulate activity: compression enforces high cooperativity, inhibiting the enzyme, whereas expansion relieves this cooperativity and activates the enzyme. We further show that all four ribonucleoside triphosphates act in symmetric pairs to tune this motion, with the pyrimidines CTP and UTP compressing the enzyme to limit further pyrimidine production, and the purines ATP and GTP expanding it to balance pyrimidine and purine pools. Together, these findings uncover a dynamic breathing mechanism for long-range allosteric communication in ATCase.
History
DepositionNov 19, 2024-
Header (metadata) releaseApr 15, 2026-
Map releaseApr 15, 2026-
UpdateMay 27, 2026-
Current statusMay 27, 2026Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_47956.png

Downloads & links

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Map

FileDownload / File: emd_47956.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.31 Å/pix.
x 360 pix.
= 471.6 Å		1.31 Å/pix.
x 360 pix.
= 471.6 Å		1.31 Å/pix.
x 360 pix.
= 471.6 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.31 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.00717
Minimum - Maximum-0.021274932 - 0.03827462
Average (Standard dev.)-0.000000022053944 (±0.00064766966)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 471.59998 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: #1

Fileemd_47956_additional_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Half map: #2

Fileemd_47956_half_map_1.map
Projections & Slices
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Histogram

Histogram (log scale)

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Half map: #1

Fileemd_47956_half_map_2.map
Projections & Slices
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Sample components

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Entire : Heterododecameric aspartate transcarbamoylase

EntireName: Heterododecameric aspartate transcarbamoylase
Components
  • Complex: Heterododecameric aspartate transcarbamoylase
    • Protein or peptide: Aspartate carbamoyltransferase regulatory chain
    • Protein or peptide: Aspartate carbamoyltransferase
  • Ligand: ZINC ION

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Supramolecule #1: Heterododecameric aspartate transcarbamoylase

SupramoleculeName: Heterododecameric aspartate transcarbamoylase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Details: Prepared in 40 mM Tris, 15 mM MgCl2, 1 mM TCEP, pH 7.5
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: Aspartate carbamoyltransferase regulatory chain

MacromoleculeName: Aspartate carbamoyltransferase regulatory chain / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 17.143625 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MTHDNKLQVE AIKRGTVIDH IPAQIGFKLL SLFKLTETDQ RITIGLNLPS GEMGRKDLIK IENTFLSEDQ VDQLALYAPQ ATVNRIDNY EVVGKSRPSL PERIDNVLVC PNSNCISHAE PVSSSFAVRK RANDIALKCK YCEKEFSHNV VLAN

UniProtKB: Aspartate carbamoyltransferase regulatory chain

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Macromolecule #2: Aspartate carbamoyltransferase

MacromoleculeName: Aspartate carbamoyltransferase / type: protein_or_peptide / ID: 2 / Number of copies: 6 / Enantiomer: LEVO / EC number: aspartate carbamoyltransferase
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 34.337105 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: ANPLYQKHII SINDLSRDDL NLVLATAAKL KANPQPELLK HKVIASCFFE ASTRTRLSFE TSMHRLGASV VGFSDSANTS LGKKGETLA DTISVISTYV DAIVMRHPQE GAARLATEFS GNVPVLNAGD GSNQHPTQTL LDLFTIQETQ GRLDNLHVAM V GDLKYGRT ...String:
ANPLYQKHII SINDLSRDDL NLVLATAAKL KANPQPELLK HKVIASCFFE ASTRTRLSFE TSMHRLGASV VGFSDSANTS LGKKGETLA DTISVISTYV DAIVMRHPQE GAARLATEFS GNVPVLNAGD GSNQHPTQTL LDLFTIQETQ GRLDNLHVAM V GDLKYGRT VHSLTQALAK FDGNRFYFIA PDALAMPQYI LDMLDEKGIA WSLHSSIEEV MAEVDILYMT RVQKERLDPS EY ANVKAQF VLRASDLHNA KANMKVLHPL PRVDEIATDV DKTPHAWYFQ QAGNGIFARQ ALLALVLNRD LVL

UniProtKB: Aspartate carbamoyltransferase

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Macromolecule #3: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 3 / Number of copies: 6 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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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: 7.5
Component:
ConcentrationFormulaName
40.0 mMC4H11NO3Tris(hydroxymethyl)aminomethane hydrochloride
15.0 mMMgCl2Magnesium chloride
1.0 mMC9H15O6PTris(2-carboxyethyl)phosphine
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Details: Mention GO.
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.6 µm
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.62 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 565000
Initial angle assignmentType: RANDOM ASSIGNMENT
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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