|Entry||Database: PDB / ID: 5u05|
|Title||Cryo-EM structure of the E. coli CTP synthase tetramer|
|Components||CTP synthaseCTP synthetase|
|Keywords||LIGASE / nucleotide metabolism / enzyme / active|
|Function/homology||CTP synthase / CTP synthase / CTP synthase GATase domain / CTP synthase activity / CTP synthase, N-terminal / CTP synthase N-terminus / 'de novo' CTP biosynthetic process / Glutamine amidotransferase / Glutamine amidotransferase type 1 domain profile. / CTP biosynthetic process ...CTP synthase / CTP synthase / CTP synthase GATase domain / CTP synthase activity / CTP synthase, N-terminal / CTP synthase N-terminus / 'de novo' CTP biosynthetic process / Glutamine amidotransferase / Glutamine amidotransferase type 1 domain profile. / CTP biosynthetic process / Glutamine amidotransferase class-I / glutamine metabolic process / Class I glutamine amidotransferase-like / P-loop containing nucleoside triphosphate hydrolase / ATP binding / identical protein binding / metal ion binding / cytosol / CTP synthase / CTP synthase|
Function and homology information
|Specimen source||Escherichia coli / / bacteria /|
|Method||Electron microscopy (7.9 Å resolution / Particle / Single particle) / Transmission electron microscopy|
|Authors||Lynch, E.M. / Hicks, D.R. / Kollman, J.M.|
|Citation||Journal: 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.
SummaryFull reportAbout validation report
|Date||Deposition: Nov 22, 2016 / Release: Apr 26, 2017|
Downloads & links
A: CTP synthase
B: CTP synthase
C: CTP synthase
D: CTP synthase
Mass: 60446.980 Da / Num. of mol.: 4 / Source: (gene. exp.) Escherichia coli / / bacteria / / Gene: pyrG, ECS88_3048 / Production host: Escherichia coli
References: UniProt:B7MLA1, UniProt:P0A7E5*PLUS, EC:22.214.171.124 (CTP synthase)
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / Reconstruction method: SINGLE PARTICLE|
|Component||Name: E. coli CTP synthase tetramer / Type: COMPLEX / Entity ID: 1 / Source: RECOMBINANT|
|Source (natural)||Organism: Escherichia coli|
|Source (recombinant)||Organism: Escherichia coli / Plasmid: pET22|
|Buffer solution||pH: 8|
|Specimen||Conc.: 0.3 mg/ml|
Details: 50 mM HEPES potassium, pH 8.0, 10 mM magnesium chloride, 0.6 mM UTP, 1.5 mM AMP-PNP, 0.2 mM GTP, 10 mM glutamine
Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
|Vitrification||Cryogen name: ETHANE|
-Electron microscopy imaging
Model: Tecnai F20 / Image courtesy: FEI Company
|Microscopy||Microscope model: FEI TECNAI F20|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: OTHER|
|Electron lens||Mode: BRIGHT FIELD|
|Image recording||Electron dose: 68 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)|
|CTF correction||Type: PHASE FLIPPING ONLY|
|Symmetry||Point symmetry: D2|
|3D reconstruction||Resolution: 7.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 6407 / Symmetry type: POINT|
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