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- EMDB-33309: E.coli phosphoribosylpyrophosphate (PRPP) synthetase type B filam... -

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Basic information

Entry
Database: EMDB / ID: EMD-33309
TitleE.coli phosphoribosylpyrophosphate (PRPP) synthetase type B filament bound with Pi
Map data
Sample
  • Organelle or cellular component: E.coli PRPP syhthetase type B filament structure complex with Pi
    • Protein or peptide: Ribose-phosphate pyrophosphokinase
  • Ligand: PHOSPHATE ION
  • Ligand: water
KeywordsAllosteric enzyme / Kinase / Transferase / Nucleotide biosynthesis / ATP-binding / Magnesium / Manganese / Metal-binding / Nucleotide-binding / BIOSYNTHETIC PROTEIN
Function / homology
Function and homology information


ribonucleoside monophosphate biosynthetic process / ribose phosphate diphosphokinase complex / ribose-phosphate diphosphokinase / ribose phosphate diphosphokinase activity / 5-phosphoribose 1-diphosphate biosynthetic process / purine nucleotide biosynthetic process / protein hexamerization / phosphate ion binding / ADP binding / kinase activity ...ribonucleoside monophosphate biosynthetic process / ribose phosphate diphosphokinase complex / ribose-phosphate diphosphokinase / ribose phosphate diphosphokinase activity / 5-phosphoribose 1-diphosphate biosynthetic process / purine nucleotide biosynthetic process / protein hexamerization / phosphate ion binding / ADP binding / kinase activity / magnesium ion binding / ATP binding / identical protein binding / cytosol / cytoplasm
Similarity search - Function
Phosphoribosyl pyrophosphate synthetase, conserved site / Phosphoribosyl pyrophosphate synthase signature. / Ribose-phosphate pyrophosphokinase, bacterial-type / Phosphoribosyl synthetase-associated domain / N-terminal domain of ribose phosphate pyrophosphokinase / Ribose-phosphate pyrophosphokinase / Ribose-phosphate pyrophosphokinase, N-terminal domain / N-terminal domain of ribose phosphate pyrophosphokinase / Phosphoribosyltransferase-like / Phosphoribosyltransferase domain
Similarity search - Domain/homology
Ribose-phosphate pyrophosphokinase
Similarity search - Component
Biological speciesEscherichia coli str. K-12 substr. MG1655 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsHu HH / Lu GM / Chang CC / Liu JL
Funding support China, 2 items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, China)No. 2021YFA0804701-4 China
National Natural Science Foundation of China (NSFC)No. 31771490 China
CitationJournal: Elife / Year: 2022
Title: Filamentation modulates allosteric regulation of PRPS.
Authors: Huan-Huan Hu / Guang-Ming Lu / Chia-Chun Chang / Yilan Li / Jiale Zhong / Chen-Jun Guo / Xian Zhou / Boqi Yin / Tianyi Zhang / Ji-Long Liu /
Abstract: Phosphoribosyl pyrophosphate (PRPP) is a key intermediate in the biosynthesis of purine and pyrimidine nucleotides, histidine, tryptophan, and cofactors NAD and NADP. Abnormal regulation of PRPP ...Phosphoribosyl pyrophosphate (PRPP) is a key intermediate in the biosynthesis of purine and pyrimidine nucleotides, histidine, tryptophan, and cofactors NAD and NADP. Abnormal regulation of PRPP synthase (PRPS) is associated with human disorders, including Arts syndrome, retinal dystrophy, and gouty arthritis. Recent studies have demonstrated that PRPS can form filamentous cytoophidia in eukaryotes. Here, we show that PRPS forms cytoophidia in prokaryotes both in vitro and in vivo. Moreover, we solve two distinct filament structures of PRPS at near-atomic resolution using Cryo-EM. The formation of the two types of filaments is controlled by the binding of different ligands. One filament type is resistant to allosteric inhibition. The structural comparison reveals conformational changes of a regulatory flexible loop, which may regulate the binding of the allosteric inhibitor and the substrate ATP. A noncanonical allosteric AMP/ADP binding site is identified to stabilize the conformation of the regulatory flexible loop. Our findings not only explore a new mechanism of PRPS regulation with structural basis, but also propose an additional layer of cell metabolism through PRPS filamentation.
History
DepositionApr 27, 2022-
Header (metadata) releaseJun 29, 2022-
Map releaseJun 29, 2022-
UpdateJul 3, 2024-
Current statusJul 3, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33309.png

Downloads & links

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Map

FileDownload / File: emd_33309.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 256 pix.
= 271.36 Å		1.06 Å/pix.
x 256 pix.
= 271.36 Å		1.06 Å/pix.
x 256 pix.
= 271.36 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.06 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.035
Minimum - Maximum-0.23506467 - 0.40432954
Average (Standard dev.)-0.000013631732 (±0.009324222)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 271.36 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_33309_msk_1.map
Projections & Slices
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Additional map: C1 symmetry of the type B filament map...

Fileemd_33309_additional_1.map
AnnotationC1 symmetry of the type B filament map to show the interface of different hexamers.The mask is not suitable for this map.
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Half map: #2

Fileemd_33309_half_map_1.map
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Half map: #1

Fileemd_33309_half_map_2.map
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Sample components

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Entire : E.coli PRPP syhthetase type B filament structure complex with Pi

EntireName: E.coli PRPP syhthetase type B filament structure complex with Pi
Components
  • Organelle or cellular component: E.coli PRPP syhthetase type B filament structure complex with Pi
    • Protein or peptide: Ribose-phosphate pyrophosphokinase
  • Ligand: PHOSPHATE ION
  • Ligand: water

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Supramolecule #1: E.coli PRPP syhthetase type B filament structure complex with Pi

SupramoleculeName: E.coli PRPP syhthetase type B filament structure complex with Pi
type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Escherichia coli str. K-12 substr. MG1655 (bacteria)

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Macromolecule #1: Ribose-phosphate pyrophosphokinase

MacromoleculeName: Ribose-phosphate pyrophosphokinase / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO / EC number: ribose-phosphate diphosphokinase
Source (natural)Organism: Escherichia coli str. K-12 substr. MG1655 (bacteria)
Molecular weightTheoretical: 35.08409 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MPDMKLFAGN ATPELAQRIA NRLYTSLGDA AVGRFSDGEV SVQINENVRG GDIFIIQSTC APTNDNLMEL VVMVDALRRA SAGRITAVI PYFGYARQDR RVRSARVPIT AKVVADFLSS VGVDRVLTVD LHAEQIQGFF DVPVDNVFGS PILLEDMLQL N LDNPIVVS ...String:
MPDMKLFAGN ATPELAQRIA NRLYTSLGDA AVGRFSDGEV SVQINENVRG GDIFIIQSTC APTNDNLMEL VVMVDALRRA SAGRITAVI PYFGYARQDR RVRSARVPIT AKVVADFLSS VGVDRVLTVD LHAEQIQGFF DVPVDNVFGS PILLEDMLQL N LDNPIVVS PDIGGVVRAR AIAKLLNDTD MAIIDKRRPR ANVSQVMHII GDVAGRDCVL VDDMIDTGGT LCKAAEALKE RG AKRVFAY ATHPIFSGNA ANNLRNSVID EVVVCDTIPL SDEIKSLPNV RTLTLSGMLA EAIRRISNEE SISAMFEHHH HHH H

UniProtKB: Ribose-phosphate pyrophosphokinase

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Macromolecule #2: PHOSPHATE ION

MacromoleculeName: PHOSPHATE ION / type: ligand / ID: 2 / Number of copies: 12 / Formula: PO4
Molecular weightTheoretical: 94.971 Da
Chemical component information

ChemComp-PO4:
PHOSPHATE ION

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Macromolecule #3: water

MacromoleculeName: water / type: ligand / ID: 3 / Number of copies: 54 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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

Concentration0.2 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
100.0 mMNaClsodium chloeide
50.0 mMNa2HPO4sodium dihydrogen phosphate anhydrous
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
Details: blot for 3.5 seconds with blot force of -1 before plunge-freezing.
DetailsThe purifide monomer or oligomer protein was incubated with Pi to form this type B filament.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 3131 / Average exposure time: 4.0 sec. / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 22500
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1186879
Startup modelType of model: OTHER
Details: The featureless cylinder was reconstructed using the relion_helix_toolbox command and applied as a reference model for 3D classification.
Final reconstructionNumber classes used: 3 / Applied symmetry - Point group: D3 (2x3 fold dihedral) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1-beta) / Number images used: 168218
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final 3D classificationNumber classes: 6 / Avg.num./class: 91698 / Software - Name: RELION
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

4s2u


Chain - Chain ID: A / Chain - Residue range: 2-309 / Chain - Source name: PDB / Chain - Initial model type: experimental model
Output model

PDB-7xn3:
E.coli phosphoribosylpyrophosphate (PRPP) synthetase type B filament bound with Pi

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