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- EMDB-20352: Tetrameric cryo-EM ArnA -

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Basic information

Entry
Database: EMDB / ID: EMD-20352
TitleTetrameric cryo-EM ArnA
Map dataLocal sharpened tetrameric cryo-EM ArnA
Sample
  • Complex: ArnA
    • Protein or peptide: Bifunctional polymyxin resistance protein ArnA
    • Protein or peptide: UDP-4-amino-4-deoxy-L-arabinose formyltransferase
KeywordsPolymyxin resistance / hexamer / tetramer / ANTIBIOTIC
Function / homology
Function and homology information


: / UDP-4-deoxy-4-formamido-beta-L-arabinopyranose biosynthetic process / UDP-glucuronic acid dehydrogenase activity / UDP-4-amino-4-deoxy-L-arabinose formyltransferase activity / UDP-glucuronic acid dehydrogenase (UDP-4-keto-hexauronic acid decarboxylating) / UDP-4-amino-4-deoxy-L-arabinose formyltransferase / UDP-D-xylose biosynthetic process / UDP-glucuronate decarboxylase activity / lipopolysaccharide biosynthetic process / lipid A biosynthetic process ...: / UDP-4-deoxy-4-formamido-beta-L-arabinopyranose biosynthetic process / UDP-glucuronic acid dehydrogenase activity / UDP-4-amino-4-deoxy-L-arabinose formyltransferase activity / UDP-glucuronic acid dehydrogenase (UDP-4-keto-hexauronic acid decarboxylating) / UDP-4-amino-4-deoxy-L-arabinose formyltransferase / UDP-D-xylose biosynthetic process / UDP-glucuronate decarboxylase activity / lipopolysaccharide biosynthetic process / lipid A biosynthetic process / NAD+ binding / response to antibiotic / protein-containing complex / identical protein binding
Similarity search - Function
Bifunctional polymyxin resistance protein, ArnA / Bifunctional polymyxin resistance protein ArnA-like, extended (e) SDRs / Formyl transferase, C-terminal / Formyl transferase, C-terminal domain / Formyl transferase-like, C-terminal domain superfamily / Formyl transferase, N-terminal / Formyl transferase / Formyl transferase, N-terminal domain superfamily / NAD-dependent epimerase/dehydratase / NAD dependent epimerase/dehydratase family / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
Bifunctional polymyxin resistance protein ArnA / Bifunctional polymyxin resistance protein ArnA / Bifunctional polymyxin resistance protein ArnA
Similarity search - Component
Biological speciesEscherichia coli DH5[alpha] (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.8 Å
AuthorsYang M / Gehring K
Funding support Canada, 1 items
OrganizationGrant numberCountry
Canadian Institutes of Health Research (CIHR)152569 Canada
CitationJournal: J Struct Biol / Year: 2019
Title: Cryo-electron microscopy structures of ArnA, a key enzyme for polymyxin resistance, revealed unexpected oligomerizations and domain movements.
Authors: Meng Yang / Yu Seby Chen / Muneyoshi Ichikawa / Daniel Calles-Garcia / Kaustuv Basu / Rayan Fakih / Khanh Huy Bui / Kalle Gehring /
Abstract: Gram-negative bacteria evade the attack of cationic antimicrobial peptides through modifying their lipid A structure in their outer membranes with 4-amino-4-deoxy-L-arabinose (Ara4N). ArnA is a ...Gram-negative bacteria evade the attack of cationic antimicrobial peptides through modifying their lipid A structure in their outer membranes with 4-amino-4-deoxy-L-arabinose (Ara4N). ArnA is a crucial enzyme in the lipid A modification pathway and its deletion abolishes the polymyxin resistance of gram-negative bacteria. Previous studies by X-ray crystallography have shown that full-length ArnA forms a three-bladed propeller-shaped hexamer. Here, the structures of ArnA determined by cryo-electron microscopy (cryo-EM) reveal that ArnA exists in two 3D architectures, hexamer and tetramer. This is the first observation of a tetrameric ArnA. The hexameric cryo-EM structure is similar to previous crystal structures but shows differences in domain movements and conformational changes. We propose that ArnA oligomeric states are in a dynamic equilibrium, where the hexamer state is energetically more favorable, and its domain movements are important for cooperating with downstream enzymes in the lipid A-Ara4N modification pathway. The results provide us with new possibilities to explore inhibitors targeting ArnA.
History
DepositionJun 26, 2019-
Header (metadata) releaseJul 31, 2019-
Map releaseJul 31, 2019-
UpdateMar 20, 2024-
Current statusMar 20, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 9
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 9
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6pik
  • Surface level: 9
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20352.png

Downloads & links

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Map

FileDownload / File: emd_20352.map.gz / Format: CCP4 / Size: 35.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationLocal sharpened tetrameric cryo-EM ArnA
Voxel sizeX=Y=Z: 1.073 Å
Density
Contour LevelBy AUTHOR: 9.0 / Movie #1: 9
Minimum - Maximum-13.386502999999999 - 24.756440000000001
Average (Standard dev.)0.3017026 (±1.8188617)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions210210210
Spacing210210210
CellA=B=C: 225.32999 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0731.0731.073
M x/y/z210210210
origin x/y/z0.0000.0000.000
length x/y/z225.330225.330225.330
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS210210210
D min/max/mean-13.38724.7560.302

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Supplemental data

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Half map: The half map of tetrameric cryo-EM ArnA

Fileemd_20352_half_map_1.map
AnnotationThe half map of tetrameric cryo-EM ArnA
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: The half map of tetrameric cryo-EM ArnA

Fileemd_20352_half_map_2.map
AnnotationThe half map of tetrameric cryo-EM ArnA
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : ArnA

EntireName: ArnA
Components
  • Complex: ArnA
    • Protein or peptide: Bifunctional polymyxin resistance protein ArnA
    • Protein or peptide: UDP-4-amino-4-deoxy-L-arabinose formyltransferase

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Supramolecule #1: ArnA

SupramoleculeName: ArnA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all / Details: Full length ArnA from E.coli
Source (natural)Organism: Escherichia coli DH5[alpha] (bacteria)
Molecular weightTheoretical: 440 KDa

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Macromolecule #1: Bifunctional polymyxin resistance protein ArnA

MacromoleculeName: Bifunctional polymyxin resistance protein ArnA / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
EC number: UDP-4-amino-4-deoxy-L-arabinose formyltransferase
Source (natural)Organism: Escherichia coli DH5[alpha] (bacteria)
Molecular weightTheoretical: 32.717416 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MKTVVFAYHD MGCLGIEALL AAGYEISAIF THTDNPGEKA FYGSVARLAA ERGIPVYAPD NVNHPLWVER IAQLSPDVIF SFYYRHLIY DEILQLAPAG AFNLHGSLLP KYRGRAPLNW VLVNGETETG VTLHRMVKRA DAGAIVAQLR IAIAPDDIAI T LHHKLCHA ...String:
MKTVVFAYHD MGCLGIEALL AAGYEISAIF THTDNPGEKA FYGSVARLAA ERGIPVYAPD NVNHPLWVER IAQLSPDVIF SFYYRHLIY DEILQLAPAG AFNLHGSLLP KYRGRAPLNW VLVNGETETG VTLHRMVKRA DAGAIVAQLR IAIAPDDIAI T LHHKLCHA ARQLLEQTLP AIKHGNILEI AQRENEATCF GRRTPDDSFL EWHKPASVLH NMVRAVADPW PGAFSYVGNQ KF TVWSSRV HPHASKAQPG SVISVAPLLI ACGDGALEIV TGQAGDGITM QGSQLAQTLG LVQ

UniProtKB: Bifunctional polymyxin resistance protein ArnA

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Macromolecule #2: UDP-4-amino-4-deoxy-L-arabinose formyltransferase

MacromoleculeName: UDP-4-amino-4-deoxy-L-arabinose formyltransferase / type: protein_or_peptide / ID: 2 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli DH5[alpha] (bacteria)
Molecular weightTheoretical: 40.057586 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MRVLILGVNG FIGNHLTERL LREDHYEVYG LDIGSDAISR FLNHPHFHFV EGDISIHSEW IEYHVKKCDV VLPLVAIATP IEYTRNPLR VFELDFEENL RIIRYCVKYR KRIIFPSTSE VYGMCSDKYF DEDHSNLIVG PVNKPRWIYS VSKQLLDRVI W AYGEKEGL ...String:
MRVLILGVNG FIGNHLTERL LREDHYEVYG LDIGSDAISR FLNHPHFHFV EGDISIHSEW IEYHVKKCDV VLPLVAIATP IEYTRNPLR VFELDFEENL RIIRYCVKYR KRIIFPSTSE VYGMCSDKYF DEDHSNLIVG PVNKPRWIYS VSKQLLDRVI W AYGEKEGL QFTLFRPFNW MGPRLDNLNA ARIGSSRAIT QLILNLVEGS PIKLIDGGKQ KRCFTDIRDG IEALYRIIEN AG NRCDGEI INIGNPENEA SIEELGEMLL ASFEKHPLRH HFPPFAGFRV VESSSYYGKG YQDVEHRKPS IRNAHRCLDW EPK IDMQET IDETLDFFLR TVDLTDKPS

UniProtKB: Bifunctional polymyxin resistance protein ArnA

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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

Concentration1.2 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClSodium chloridesodium chloride
20.0 mMC4H11NO3Tris
1.0 mMC9H15O6PHClTCEP-HCl
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Average electron dose: 35.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

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

4wkg

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 7.8 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 38127
FSC plot (resolution estimation)

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