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- EMDB-16647: Cryo-EM structure of RNase J from Helicobacter pylori -

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Basic information

Entry
Database: EMDB / ID: EMD-16647
TitleCryo-EM structure of RNase J from Helicobacter pylori
Map data
Sample
  • Complex: Helicobacter pylori RNase J
    • Protein or peptide: Ribonuclease J
Keywordsribonuclease / RNase J / degradosome / Helicobacter pylori / RNA BINDING PROTEIN
Function / homology
Function and homology information


5'-3' RNA exonuclease activity / RNA endonuclease activity / mRNA processing / rRNA processing / Hydrolases; Acting on ester bonds / RNA binding / zinc ion binding / cytoplasm
Similarity search - Function
Ribonuclease J, conserved site / Uncharacterized protein family UPF0036 signature. / Ribonuclease J, bacteria / Ribonuclease J, C-terminal / Ribonuclease J C-terminal domain / Ribonuclease J / Ribonuclease J, domain 2 / Zn-dependent metallo-hydrolase, RNA specificity domain / Zn-dependent metallo-hydrolase RNA specificity domain / Metallo-beta-lactamase superfamily ...Ribonuclease J, conserved site / Uncharacterized protein family UPF0036 signature. / Ribonuclease J, bacteria / Ribonuclease J, C-terminal / Ribonuclease J C-terminal domain / Ribonuclease J / Ribonuclease J, domain 2 / Zn-dependent metallo-hydrolase, RNA specificity domain / Zn-dependent metallo-hydrolase RNA specificity domain / Metallo-beta-lactamase superfamily / Metallo-beta-lactamase superfamily / Metallo-beta-lactamase / Ribonuclease Z/Hydroxyacylglutathione hydrolase-like
Similarity search - Domain/homology
Biological speciesHelicobacter pylori (bacteria) / Helicobacter pylori B128 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsLulla A / Luisi BF
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Wellcome Trust200873/Z/16/Z United Kingdom
CitationJournal: Nat Commun / Year: 2023
Title: Acetylation regulates the oligomerization state and activity of RNase J, the Helicobacter pylori major ribonuclease.
Authors: Alejandro Tejada-Arranz / Aleksei Lulla / Maxime Bouilloux-Lafont / Evelyne Turlin / Xue-Yuan Pei / Thibaut Douché / Mariette Matondo / Allison H Williams / Bertrand Raynal / Ben F Luisi / Hilde De Reuse /
Abstract: In the gastric pathogen Helicobacter pylori, post-transcriptional regulation relies strongly on the activity of the essential ribonuclease RNase J. Here, we elucidated the crystal and cryo-EM ...In the gastric pathogen Helicobacter pylori, post-transcriptional regulation relies strongly on the activity of the essential ribonuclease RNase J. Here, we elucidated the crystal and cryo-EM structures of RNase J and determined that it assembles into dimers and tetramers in vitro. We found that RNase J extracted from H. pylori is acetylated on multiple lysine residues. Alanine substitution of several of these residues impacts on H. pylori morphology, and thus on RNase J function in vivo. Mutations of Lysine 649 modulates RNase J oligomerization in vitro, which in turn influences ribonuclease activity in vitro. Our structural analyses of RNase J reveal loops that gate access to the active site and rationalizes how acetylation state of K649 can influence activity. We propose acetylation as a regulatory level controlling the activity of RNase J and its potential cooperation with other enzymes of RNA metabolism in H. pylori.
History
DepositionFeb 5, 2023-
Header (metadata) releaseDec 27, 2023-
Map releaseDec 27, 2023-
UpdateDec 27, 2023-
Current statusDec 27, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16647.png

Downloads & links

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Map

FileDownload / File: emd_16647.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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AxesZ (Sec.)Y (Row.)X (Col.)
1.11 Å/pix.
x 256 pix.
= 283.136 Å		1.11 Å/pix.
x 256 pix.
= 283.136 Å		1.11 Å/pix.
x 256 pix.
= 283.136 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.106 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.184
Minimum - Maximum-0.27704468 - 0.779917
Average (Standard dev.)0.0019870508 (±0.035276823)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 283.136 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_16647_msk_1.map
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Half map: #2

Fileemd_16647_half_map_1.map
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Half map: #1

Fileemd_16647_half_map_2.map
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Sample components

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Entire : Helicobacter pylori RNase J

EntireName: Helicobacter pylori RNase J
Components
  • Complex: Helicobacter pylori RNase J
    • Protein or peptide: Ribonuclease J

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Supramolecule #1: Helicobacter pylori RNase J

SupramoleculeName: Helicobacter pylori RNase J / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Helicobacter pylori (bacteria) / Strain: B128
Molecular weightTheoretical: 316 KDa

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Macromolecule #1: Ribonuclease J

MacromoleculeName: Ribonuclease J / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO / EC number: Hydrolases; Acting on ester bonds
Source (natural)Organism: Helicobacter pylori B128 (bacteria)
Molecular weightTheoretical: 79.19693 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: GSGTDNNHYE NNESNENSSE NSKVDEARAG AFERFTNRKK RFRENAQKNG ESSHHEAPSH HKKEHRPNKK PNNHHKQKHA KTRNYAKEE LDSNKVEGVT EILHVNERGT LGFHKELKKG VETNNKIQVE HLNPHYKMNL NSKASVKITP LGGLGEIGGN M MVIETPKS ...String:
GSGTDNNHYE NNESNENSSE NSKVDEARAG AFERFTNRKK RFRENAQKNG ESSHHEAPSH HKKEHRPNKK PNNHHKQKHA KTRNYAKEE LDSNKVEGVT EILHVNERGT LGFHKELKKG VETNNKIQVE HLNPHYKMNL NSKASVKITP LGGLGEIGGN M MVIETPKS AIVIDAGMSF PKEGLFGVDI LIPDFSYLHQ IKDKIAGIII THAHEDHIGA TPYLFKELQF PLYGTPLSLG LI GSKFDEH GLKKYRSYFK IVEKRCPISV GEFIIEWIHI THSIIDSSAL AIQTKAGTII HTGDFKIDHT PVDNLPTDLY RLA HYGEKG VMLLLSDSTN SHKSGTTPSE STIAPAFDTL FKEAQGRVIM STFSSNIHRV YQAIQYGIKY NRKIAVIGRS MEKN LDIAR ELGYIHLPYQ SFIEANEVAK YPDNEVLIVT TGSQGETMSA LYRMATDEHR HISIKPNDLV IISAKAIPGN EASVS AVLN FLIKKEAKVA YQEFDNIHVS GHAAQEEQKL MLRLIKPKFF LPVHGEYNHV ARHKQTAISC GVPEKNIYLM EDGDQV EVG PAFIKKVGTI KSGKSYVDNQ SNLSIDTSIV QQREEVASAG VFAATIFVNK NKQALLESSQ FSSLGLVGFK DEKPLIK EI QGGLEMLLKS SNAEILNNPK KLEDHTRNFI RKALFKKFRK YPAIICHAHS FGSSPHHHHH HHH

UniProtKB: Ribonuclease J

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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

Concentration0.3 mg/mL
BufferpH: 7.2
Component:
ConcentrationFormulaName
20.0 mMNa2HPO4sodium phosphate
500.0 mMNaClSodium chloridesodium chloride
0.5 mMTCEP
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Support film - Material: CARBON / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.029 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated defocus min: 1.2 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.2 µm
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Average electron dose: 40.58 e/Å2
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

7pcr

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 34061
FSC plot (resolution estimation)

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