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- EMDB-15784: CryoEM structure of bacterial RapZ.GlmZ complex central to the co... -

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Basic information

Entry
Database: EMDB / ID: EMD-15784
TitleCryoEM structure of bacterial RapZ.GlmZ complex central to the control of cell envelope biogenesis
Map data
Sample
  • Complex: RapZ tetramer, GlmZ stem loops I & II
    • Protein or peptide: RNase adapter protein RapZ
    • RNA: GlmZ small regulatory RNA
Function / homology
Function and homology information


RNA destabilization / carbohydrate derivative binding / protein homotetramerization / molecular adaptor activity / GTP binding / protein-containing complex / RNA binding / ATP binding / identical protein binding
Similarity search - Function
RapZ-like family / RapZ-like N-terminal domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
RNase adapter protein RapZ
Similarity search - Component
Biological speciesEscherichia coli (E. coli) / Escherichia coli K-12 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.28 Å
AuthorsIslam MS / Hardwick HW / Chirgadze DY / Luisi BF
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Wellcome Trust200873/Z/16/Z United Kingdom
Citation
Journal: EMBO J / Year: 2023
Title: Structure of a bacterial ribonucleoprotein complex central to the control of cell envelope biogenesis.
Authors: Md Saiful Islam / Steven W Hardwick / Laura Quell / Svetlana Durica-Mitic / Dimitri Y Chirgadze / Boris Görke / Ben F Luisi /
Abstract: Biogenesis of the essential precursor of the bacterial cell envelope, glucosamine-6-phosphate (GlcN6P), is controlled by intricate post-transcriptional networks mediated by GlmZ, a small regulatory ...Biogenesis of the essential precursor of the bacterial cell envelope, glucosamine-6-phosphate (GlcN6P), is controlled by intricate post-transcriptional networks mediated by GlmZ, a small regulatory RNA (sRNA). GlmZ stimulates translation of the mRNA encoding GlcN6P synthtase in Escherichia coli, but when bound by RapZ protein, the sRNA becomes inactivated through cleavage by the endoribonuclease RNase E. Here, we report the cryoEM structure of the RapZ:GlmZ complex, revealing a complementary match of the RapZ tetrameric quaternary structure to structural repeats in the sRNA. The nucleic acid is contacted by RapZ mostly through a highly conserved domain that shares an evolutionary relationship with phosphofructokinase and suggests links between metabolism and riboregulation. We also present the structure of a precleavage intermediate formed between the binary RapZ:GlmZ complex and RNase E that reveals how GlmZ is presented and recognised by the enzyme. The structures provide a framework for understanding how other encounter complexes might guide recognition and action of endoribonucleases on target transcripts, and how structured substrates in polycistronic precursors may be recognised for processing by RNase E.
#1: Journal: Biorxiv / Year: 2022
Title: Structure of a bacterial ribonucleoprotein complex central to the control of cell envelope biogenesis
Authors: Islam MS / Hardwick SW / Quell L / Chirgadze DY / Gorke B / Luisi BF
History
DepositionSep 7, 2022-
Header (metadata) releaseOct 5, 2022-
Map releaseOct 5, 2022-
UpdateJan 25, 2023-
Current statusJan 25, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15784.png

Downloads & links

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Map

FileDownload / File: emd_15784.map.gz / Format: CCP4 / Size: 371.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
0.65 Å/pix.
x 460 pix.
= 299.92 Å		0.65 Å/pix.
x 460 pix.
= 299.92 Å		0.65 Å/pix.
x 460 pix.
= 299.92 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.652 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.04
Minimum - Maximum-0.16635635 - 0.2576606
Average (Standard dev.)0.00010025135 (±0.007156773)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions460460460
Spacing460460460
CellA=B=C: 299.92 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_15784_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_15784_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : RapZ tetramer, GlmZ stem loops I & II

EntireName: RapZ tetramer, GlmZ stem loops I & II
Components
  • Complex: RapZ tetramer, GlmZ stem loops I & II
    • Protein or peptide: RNase adapter protein RapZ
    • RNA: GlmZ small regulatory RNA

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Supramolecule #1: RapZ tetramer, GlmZ stem loops I & II

SupramoleculeName: RapZ tetramer, GlmZ stem loops I & II / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: all / Details: In vitro reconstituted RapZ and GlmZ complex
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: RNase adapter protein RapZ

MacromoleculeName: RNase adapter protein RapZ / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli K-12 (bacteria) / Strain: K12
Molecular weightTheoretical: 32.53832 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MVLMIVSGRS GSGKSVALRA LEDMGFYCVD NLPVVLLPDL ARTLADREIS AAVSIDVRNM PESPEIFEQA MSNLPDAFSP QLLFLDADR NTLIRRYSDT RRLHPLSSKN LSLESAIDKE SDLLEPLRSR ADLIVDTSEM SVHELAEMLR TRLLGKRERE L TMVFESFG ...String:
MVLMIVSGRS GSGKSVALRA LEDMGFYCVD NLPVVLLPDL ARTLADREIS AAVSIDVRNM PESPEIFEQA MSNLPDAFSP QLLFLDADR NTLIRRYSDT RRLHPLSSKN LSLESAIDKE SDLLEPLRSR ADLIVDTSEM SVHELAEMLR TRLLGKRERE L TMVFESFG FKHGIPIDAD YVFDVRFLPN PHWDPKLRPM TGLDKPVAAF LDRHTEVHNF IYQTRSYLEL WLPMLETNNR SY LTVAIGC TGGKHRSVYI AEQLADYFRS RGKNVQSRHR TLEKRKP

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Macromolecule #2: GlmZ small regulatory RNA

MacromoleculeName: GlmZ small regulatory RNA / type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: Escherichia coli K-12 (bacteria)
Molecular weightTheoretical: 66.061859 KDa
SequenceString: GUAGAUGCUC AUUCCAUCUC UUAUGUUCGC CUUAGUGCCU CAUAAACUCC GGAAUGACGC AGAGCCGUUU ACGGUGCUUA UCGUCCACU GACAGAUGUC GCUUAUGCCU CAUCAGACAC CAUGGACACA ACGUUGAGUG AAGCACCCAC UUGUUGUCAU A CAGACCUG ...String:
GUAGAUGCUC AUUCCAUCUC UUAUGUUCGC CUUAGUGCCU CAUAAACUCC GGAAUGACGC AGAGCCGUUU ACGGUGCUUA UCGUCCACU GACAGAUGUC GCUUAUGCCU CAUCAGACAC CAUGGACACA ACGUUGAGUG AAGCACCCAC UUGUUGUCAU A CAGACCUG UUUUAACGCC UGCUCCGUUA AUAAGAGCAG GCGUUUUUU

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation state3D array

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Sample preparation

Concentration5 mg/mL
BufferpH: 7.5 / Details: 25 mM HEPES pH 7.5, 300 KCl, and 1 mM MgCl2
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 120 sec.
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: DIFFRACTION / Nominal defocus max: 0.3 µm / Nominal defocus min: 0.1 µm
Sample stageCooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 47.3 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 286172
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

5o5q

Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.28 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 27551

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