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- EMDB-20354: cryoEM structure of the substrate-bound human CTP synthase 2 filament -

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Basic information

Entry
Database: EMDB / ID: EMD-20354
TitlecryoEM structure of the substrate-bound human CTP synthase 2 filament
Map data
Sample
  • Complex: substrate-bound CTP synthase 2 filament
    • Protein or peptide: CTP synthase 2CTP synthetase
  • Ligand: URIDINE 5'-TRIPHOSPHATE
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
Function / homology
Function and homology information


CTP synthase activity / cytoophidium / CTP synthase (glutamine hydrolysing) / 'de novo' CTP biosynthetic process / pyrimidine nucleotide metabolic process / pyrimidine nucleobase biosynthetic process / CTP biosynthetic process / Interconversion of nucleotide di- and triphosphates / glutamine metabolic process / ATP binding ...CTP synthase activity / cytoophidium / CTP synthase (glutamine hydrolysing) / 'de novo' CTP biosynthetic process / pyrimidine nucleotide metabolic process / pyrimidine nucleobase biosynthetic process / CTP biosynthetic process / Interconversion of nucleotide di- and triphosphates / glutamine metabolic process / ATP binding / identical protein binding / cytosol / cytoplasm
Similarity search - Function
CTP synthase N-terminus / CTP synthase GATase domain / CTP synthase, N-terminal / CTP synthase / Glutamine amidotransferase type 1 domain profile. / Glutamine amidotransferase class-I / Glutamine amidotransferase / Class I glutamine amidotransferase-like / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.5 Å
AuthorsLynch EM / Kollman JM
CitationJournal: Nat Struct Mol Biol / Year: 2020
Title: Coupled structural transitions enable highly cooperative regulation of human CTPS2 filaments.
Authors: Eric M Lynch / Justin M Kollman /
Abstract: Many enzymes assemble into defined oligomers, providing a mechanism for cooperatively regulating activity. Recent studies have described a mode of regulation in which enzyme activity is modulated by ...Many enzymes assemble into defined oligomers, providing a mechanism for cooperatively regulating activity. Recent studies have described a mode of regulation in which enzyme activity is modulated by polymerization into large-scale filaments. Here we describe an ultrasensitive form of polymerization-based regulation employed by human CTP synthase 2 (CTPS2). Cryo-EM structures reveal that CTPS2 filaments dynamically switch between active and inactive forms in response to changes in substrate and product levels. Linking the conformational state of many CTPS2 subunits in a filament results in highly cooperative regulation, greatly exceeding the limits of cooperativity for the CTPS2 tetramer alone. The structures reveal a link between conformation and control of ammonia channeling between the enzyme's active sites, and explain differences in regulation of human CTPS isoforms. This filament-based mechanism of enhanced cooperativity demonstrates how the widespread phenomenon of enzyme polymerization can be adapted to achieve different regulatory outcomes.
History
DepositionJun 28, 2019-
Header (metadata) releaseJul 31, 2019-
Map releaseDec 25, 2019-
UpdateJan 22, 2020-
Current statusJan 22, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4.4
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 4.4
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6pk4
  • Surface level: 4.4
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6pk4
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_20354.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.05 Å/pix.
x 300 pix.
= 315. Å
1.05 Å/pix.
x 300 pix.
= 315. Å
1.05 Å/pix.
x 300 pix.
= 315. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.05 Å
Density
Contour LevelBy AUTHOR: 4.4 / Movie #1: 4.4
Minimum - Maximum-8.81555 - 15.92871
Average (Standard dev.)0.023350943 (±0.77334726)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 315.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.051.051.05
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z315.000315.000315.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-200-200-200
NX/NY/NZ401401401
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-8.81615.9290.023

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Supplemental data

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Sample components

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Entire : substrate-bound CTP synthase 2 filament

EntireName: substrate-bound CTP synthase 2 filament
Components
  • Complex: substrate-bound CTP synthase 2 filament
    • Protein or peptide: CTP synthase 2CTP synthetase
  • Ligand: URIDINE 5'-TRIPHOSPHATE
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE

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Supramolecule #1: substrate-bound CTP synthase 2 filament

SupramoleculeName: substrate-bound CTP synthase 2 filament / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Saccharomyces cerevisiae (baker's yeast)

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Macromolecule #1: CTP synthase 2

MacromoleculeName: CTP synthase 2 / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO / EC number: CTP synthase (glutamine hydrolysing)
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 65.75943 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (baker's yeast)
SequenceString: MKYILVTGGV ISGIGKGIIA SSIGTILKSC GLRVTAIKID PYINIDAGTF SPYEHGEVFV LNDGGEVDLD LGNYERFLDI NLYKDNNIT TGKIYQHVIN KERRGDYLGK TVQVVPHITD AVQEWVMNQA KVPVDGNKEE PQICVIELGG TIGDIEGMPF V EAFRQFQF ...String:
MKYILVTGGV ISGIGKGIIA SSIGTILKSC GLRVTAIKID PYINIDAGTF SPYEHGEVFV LNDGGEVDLD LGNYERFLDI NLYKDNNIT TGKIYQHVIN KERRGDYLGK TVQVVPHITD AVQEWVMNQA KVPVDGNKEE PQICVIELGG TIGDIEGMPF V EAFRQFQF KAKRENFCNI HVSLVPQLSA TGEQKTKPTQ NSVRALRGLG LSPDLIVCRS STPIEMAVKE KISMFCHVNP EQ VICIHDV SSTYRVPVLL EEQSIVKYFK ERLHLPIGDS ASNLLFKWRN MADRYERLQK ICSIALVGKY TKLRDCYASV FKA LEHSAL AINHKLNLMY IDSIDLEKIT ETEDPVKFHE AWQKLCKADG ILVPGGFGIR GTLGKLQAIS WARTKKIPFL GVCL GMQLA VIEFARNCLN LKDADSTEFR PNAPVPLVID MPEHNPGNLG GTMRLGIRRT VFKTENSILR KLYGDVPFIE ERHRH RFEV NPNLIKQFEQ NDLSFVGQDV DGDRMEIIEL ANHPYFVGVQ FHPEFSSRPM KPSPPYLGLL LAATGNLNAY LQQGCK LSS SDRYSDASDD SFSEPRIAEL EIS

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Macromolecule #2: URIDINE 5'-TRIPHOSPHATE

MacromoleculeName: URIDINE 5'-TRIPHOSPHATE / type: ligand / ID: 2 / Number of copies: 4 / Formula: UTP
Molecular weightTheoretical: 484.141 Da
Chemical component information

ChemComp-UTP:
URIDINE 5'-TRIPHOSPHATE / UTP*YM / Uridine triphosphate

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Macromolecule #3: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 4 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM / Adenosine triphosphate

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7.9
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 90.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: Gctf
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cisTEM
Final reconstructionApplied symmetry - Point group: D2 (2x2 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cisTEM / Number images used: 53964

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