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- EMDB-8474: Cryo-EM structure of the human CTP synthase filament -

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Basic information

Entry
Database: EMDB / ID: 8474
TitleCryo-EM structure of the human CTP synthase filament
Map dataHuman CTP synthase filament
Samplehuman CTP synthase 1 filament
  • CTP synthase 1
  • (ligand) x 2
Function/homologyCTP synthase / CTP synthase activity / CTP synthase GATase domain / CTP synthase, N-terminal / CTP synthase / CTP synthase N-terminus / 'de novo' CTP biosynthetic process / Glutamine amidotransferase type 1 domain profile. / Glutamine amidotransferase / B cell proliferation ...CTP synthase / CTP synthase activity / CTP synthase GATase domain / CTP synthase, N-terminal / CTP synthase / CTP synthase N-terminus / 'de novo' CTP biosynthetic process / Glutamine amidotransferase type 1 domain profile. / Glutamine amidotransferase / B cell proliferation / CTP biosynthetic process / Glutamine amidotransferase class-I / nucleobase-containing compound metabolic process / Interconversion of nucleotide di- and triphosphates / glutamine metabolic process / T cell proliferation / nucleobase-containing small molecule interconversion / Class I glutamine amidotransferase-like / response to drug / P-loop containing nucleoside triphosphate hydrolase / membrane / ATP binding / identical protein binding / cytosol / CTP synthase 1
Function and homology information
SourceHomo sapiens / / human
MethodCryo EM / helical reconstruction / 6.1 Å resolution
AuthorsLynch EM / Kollman JM
CitationJournal: Nat. Struct. Mol. Biol. / Year: 2017
Title: Human CTP synthase filament structure reveals the active enzyme conformation.
Authors: Eric M Lynch / Derrick R Hicks / Matthew Shepherd / James A Endrizzi / Allison Maker / Jesse M Hansen / Rachael M Barry / Zemer Gitai / Enoch P Baldwin / Justin M Kollman
Abstract: The universally conserved enzyme CTP synthase (CTPS) forms filaments in bacteria and eukaryotes. In bacteria, polymerization inhibits CTPS activity and is required for nucleotide homeostasis. Here we ...The universally conserved enzyme CTP synthase (CTPS) forms filaments in bacteria and eukaryotes. In bacteria, polymerization inhibits CTPS activity and is required for nucleotide homeostasis. Here we show that for human CTPS, polymerization increases catalytic activity. The cryo-EM structures of bacterial and human CTPS filaments differ considerably in overall architecture and in the conformation of the CTPS protomer, explaining the divergent consequences of polymerization on activity. The structure of human CTPS filament, the first structure of the full-length human enzyme, reveals a novel active conformation. The filament structures elucidate allosteric mechanisms of assembly and regulation that rely on a conserved conformational equilibrium. The findings may provide a mechanism for increasing human CTPS activity in response to metabolic state and challenge the assumption that metabolic filaments are generally storage forms of inactive enzymes. Allosteric regulation of CTPS polymerization by ligands likely represents a fundamental mechanism underlying assembly of other metabolic filaments.
Validation ReportPDB-ID: 5u03

SummaryFull reportAbout validation report
DateDeposition: Nov 22, 2016 / Header (metadata) release: Apr 26, 2017 / Map release: Apr 26, 2017 / Last update: Jun 21, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3.8
  • Imaged by UCSF CHIMERA
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 3.8
  • Imaged by UCSF CHIMERA
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  • Surface view with fitted model
  • Atomic models: : PDB-5u03
  • Surface level: 3.8
  • Imaged by UCSF CHIMERA
  • Download
3D viewer
Supplemental images

Downloads & links

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Map

Fileemd_8474.map.gz (map file in CCP4 format, 226493 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
384 pix
1.26 Å/pix.
= 483.84 Å
384 pix
1.26 Å/pix.
= 483.84 Å
384 pix
1.26 Å/pix.
= 483.84 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Voxel sizeX=Y=Z: 1.26 Å
Density
Contour Level:3.8 (by author), 3.8 (movie #1):
Minimum - Maximum-8.421113 - 15.893713
Average (Standard dev.)1.69333E-10 (0.6589945)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions384384384
Origin-192-192-192
Limit191191191
Spacing384384384
CellA=B=C: 483.84 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.261.261.26
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z483.840483.840483.840
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS-192-192-192
NC/NR/NS384384384
D min/max/mean-8.42115.8940.000

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Supplemental data

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Sample components

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Entire human CTP synthase 1 filament

EntireName: human CTP synthase 1 filament
Details: hCTPS1 filaments assembled in the presence of UTP, ATP, and GTP
Number of components: 4

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Component #1: protein, human CTP synthase 1 filament

ProteinName: human CTP synthase 1 filament
Details: hCTPS1 filaments assembled in the presence of UTP, ATP, and GTP
Recombinant expression: No
SourceSpecies: Homo sapiens / / human
Source (engineered)Expression System: Saccharomyces cerevisiae / / yeast / / Vector: pDO105-hCTPS1

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Component #2: protein, CTP synthase 1

ProteinName: CTP synthase 1 / Recombinant expression: No
MassTheoretical: 66.777297 kDa
Source (engineered)Expression System: Homo sapiens / / human

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Component #3: ligand, ADENOSINE-5'-TRIPHOSPHATE

LigandName: ADENOSINE-5'-TRIPHOSPHATE / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 0.507181 kDa

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Component #4: ligand, URIDINE 5'-TRIPHOSPHATE

LigandName: URIDINE 5'-TRIPHOSPHATE / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 0.484141 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: filament / Method: Cryo EM
Helical parametersAxial symmetry: C1 (asymmetric) / Delta z: 104.1 Å / Delta phi: 60.6 deg.
Sample solutionSpecimen conc.: 0.3 mg/ml / pH: 7.9
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company
ImagingMicroscope: FEI TECNAI F20
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 45 e/Å2 / Illumination mode: OTHER
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 (4k x 4k)

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Image processing

ProcessingMethod: helical reconstruction
3D reconstructionSoftware: SPIDER, hsearch_lorentz, ctffind / Resolution: 6.1 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Output model

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