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- EMDB-36455: Cryo-EM structure of a Legionella effector complexed with actin a... -

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Basic information

Entry
Database: EMDB / ID: EMD-36455
TitleCryo-EM structure of a Legionella effector complexed with actin and ATP
Map data
Sample
  • Complex: Cryo-EM structure of a Legionella effector complexed with actin and ATP
    • Protein or peptide: Substrate of the Dot/Icm secretion system
    • Protein or peptide: Actin, alpha skeletal muscle
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: CALCIUM ION
KeywordsAMPylation / TOXIN
Function / homology
Function and homology information


cytoskeletal motor activator activity / tropomyosin binding / myosin heavy chain binding / mesenchyme migration / troponin I binding / filamentous actin / actin filament bundle / skeletal muscle thin filament assembly / actin filament bundle assembly / striated muscle thin filament ...cytoskeletal motor activator activity / tropomyosin binding / myosin heavy chain binding / mesenchyme migration / troponin I binding / filamentous actin / actin filament bundle / skeletal muscle thin filament assembly / actin filament bundle assembly / striated muscle thin filament / skeletal muscle myofibril / actin monomer binding / stress fiber / skeletal muscle fiber development / titin binding / actin filament polymerization / filopodium / actin filament / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / calcium-dependent protein binding / lamellipodium / cell body / hydrolase activity / protein domain specific binding / calcium ion binding / positive regulation of gene expression / magnesium ion binding / ATP binding / identical protein binding / cytoplasm
Similarity search - Function
Actins signature 1. / Actin, conserved site / Actins signature 2. / Actin/actin-like conserved site / Actins and actin-related proteins signature. / Actin / Actin family / Actin / ATPase, nucleotide binding domain
Similarity search - Domain/homology
Substrate of the Dot/Icm secretion system / Actin, alpha skeletal muscle
Similarity search - Component
Biological speciesLegionella sainthelensi (bacteria) / Oryctolagus cuniculus (rabbit)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.04 Å
AuthorsZhou XT / Wang XF / Tan JX / Zhu YQ
Funding support China, 4 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)81925024 China
National Natural Science Foundation of China (NSFC)21974002 China
National Natural Science Foundation of China (NSFC)22174003 China
Ministry of Science and Technology (MoST, China)2017YFA0503900 China
CitationJournal: Nature / Year: 2024
Title: Legionella effector LnaB is a phosphoryl-AMPylase that impairs phosphosignalling.
Authors: Ting Wang / Xiaonan Song / Jiaxing Tan / Wei Xian / Xingtong Zhou / Mingru Yu / Xiaofei Wang / Yan Xu / Ting Wu / Keke Yuan / Yu Ran / Bing Yang / Gaofeng Fan / Xiaoyun Liu / Yan Zhou / Yongqun Zhu /
Abstract: AMPylation is a post-translational modification in which AMP is added to the amino acid side chains of proteins. Here we show that, with ATP as the ligand and actin as the host activator, the ...AMPylation is a post-translational modification in which AMP is added to the amino acid side chains of proteins. Here we show that, with ATP as the ligand and actin as the host activator, the effector protein LnaB of Legionella pneumophila exhibits AMPylase activity towards the phosphoryl group of phosphoribose on PR-Ub that is generated by the SidE family of effectors, and deubiquitinases DupA and DupB in an E1- and E2-independent ubiquitination process. The product of LnaB is further hydrolysed by an ADP-ribosylhydrolase, MavL, to Ub, thereby preventing the accumulation of PR-Ub and ADPR-Ub and protecting canonical ubiquitination in host cells. LnaB represents a large family of AMPylases that adopt a common structural fold, distinct from those of the previously known AMPylases, and LnaB homologues are found in more than 20 species of bacterial pathogens. Moreover, LnaB also exhibits robust phosphoryl AMPylase activity towards phosphorylated residues and produces unique ADPylation modifications in proteins. During infection, LnaB AMPylates the conserved phosphorylated tyrosine residues in the activation loop of the Src family of kinases, which dampens downstream phosphorylation signalling in the host. Structural studies reveal the actin-dependent activation and catalytic mechanisms of the LnaB family of AMPylases. This study identifies, to our knowledge, an unprecedented molecular regulation mechanism in bacterial pathogenesis and protein phosphorylation.
History
DepositionJun 7, 2023-
Header (metadata) releaseMay 1, 2024-
Map releaseMay 1, 2024-
UpdateJul 24, 2024-
Current statusJul 24, 2024Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_36455.png

Downloads & links

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Map

FileDownload / File: emd_36455.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.93 Å/pix.
x 256 pix.
= 238.08 Å		0.93 Å/pix.
x 256 pix.
= 238.08 Å		0.93 Å/pix.
x 256 pix.
= 238.08 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.93 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.48
Minimum - Maximum-2.625096 - 4.4764085
Average (Standard dev.)-0.0014355241 (±0.07787997)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 238.08 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_36455_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_36455_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Cryo-EM structure of a Legionella effector complexed with actin a...

EntireName: Cryo-EM structure of a Legionella effector complexed with actin and ATP
Components
  • Complex: Cryo-EM structure of a Legionella effector complexed with actin and ATP
    • Protein or peptide: Substrate of the Dot/Icm secretion system
    • Protein or peptide: Actin, alpha skeletal muscle
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: CALCIUM ION

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Supramolecule #1: Cryo-EM structure of a Legionella effector complexed with actin a...

SupramoleculeName: Cryo-EM structure of a Legionella effector complexed with actin and ATP
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Legionella sainthelensi (bacteria)

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Macromolecule #1: Substrate of the Dot/Icm secretion system

MacromoleculeName: Substrate of the Dot/Icm secretion system / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Legionella sainthelensi (bacteria)
Molecular weightTheoretical: 50.352203 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString: MSYTKIESSL LLALDYPKLD ESDFILLLTK FLEKKLGNND NYPTFSKQIQ KYYLEQEYKK AIENILRLCQ ENETLLGTNL VQRLITKSS QVTSNPKDNE SRRFYEVLYA EHLESILRKD FDCSIFDELN EAYNEVRPEY TVNDLTKINT FEEARKLILA F VMLNDNVE ...String:
MSYTKIESSL LLALDYPKLD ESDFILLLTK FLEKKLGNND NYPTFSKQIQ KYYLEQEYKK AIENILRLCQ ENETLLGTNL VQRLITKSS QVTSNPKDNE SRRFYEVLYA EHLESILRKD FDCSIFDELN EAYNEVRPEY TVNDLTKINT FEEARKLILA F VMLNDNVE LGLKAQSAIY QKKDRSREEL GQVLTANPGI MKPNSPNFAD NTVPIKKIDK IAIDEKKAGG YSKTNPQVPF VA SLSGTTY SLVVVLQKYM DKHKTDPNLE KKINNIVMLW TSAYIKDGYA SYKEVIDIFK DAHIQSIFAR ANIKLDYAII DDT DHEFHR AQEYTQGIAT KAMMHQELVQ KVQEKSMREE KQKQMLLFCG VLVNQLNQDP KSKEKYPEQL ESLNALYKNW NAGT IKNQE FKKECTQVCT EIQIKEQNTP LEHHHHHH

UniProtKB: Substrate of the Dot/Icm secretion system

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Macromolecule #2: Actin, alpha skeletal muscle

MacromoleculeName: Actin, alpha skeletal muscle / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
EC number: Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement
Source (natural)Organism: Oryctolagus cuniculus (rabbit)
Molecular weightTheoretical: 42.096953 KDa
SequenceString: MCDEDETTAL VCDNGSGLVK AGFAGDDAPR AVFPSIVGRP RHQGVMVGMG QKDSYVGDEA QSKRGILTLK YPIEHGIITN WDDMEKIWH HTFYNELRVA PEEHPTLLTE APLNPKANRE KMTQIMFETF NVPAMYVAIQ AVLSLYASGR TTGIVLDSGD G VTHNVPIY ...String:
MCDEDETTAL VCDNGSGLVK AGFAGDDAPR AVFPSIVGRP RHQGVMVGMG QKDSYVGDEA QSKRGILTLK YPIEHGIITN WDDMEKIWH HTFYNELRVA PEEHPTLLTE APLNPKANRE KMTQIMFETF NVPAMYVAIQ AVLSLYASGR TTGIVLDSGD G VTHNVPIY EGYALPHAIM RLDLAGRDLT DYLMKILTER GYSFVTTAER EIVRDIKEKL CYVALDFENE MATAASSSSL EK SYELPDG QVITIGNERF RCPETLFQPS FIGMESAGIH ETTYNSIMKC DIDIRKDLYA NNVMSGGTTM YPGIADRMQK EIT ALAPST MKIKIIAPPE RKYSVWIGGS ILASLSTFQQ MWITKQEYDE AGPSIVHRKC F

UniProtKB: Actin, alpha skeletal muscle

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Macromolecule #3: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 2 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

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

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #5: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 5 / Number of copies: 1 / Formula: CA
Molecular weightTheoretical: 40.078 Da

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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

BufferpH: 8
GridModel: Quantifoil R0.6/1 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 180 sec.
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.6 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.04 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 171800
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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Atomic model buiding 1

Initial modelChain - Source name: Other / Chain - Initial model type: in silico model / Details: ModelAngelo built model
Output model

PDB-8jo4:
Cryo-EM structure of a Legionella effector complexed with actin and ATP

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