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- EMDB-45167: Cryo-EM structure of E. coli AmpG -

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Basic information

Entry
Database: EMDB / ID: EMD-45167
TitleCryo-EM structure of E. coli AmpG
Map data
Sample
  • Complex: AmpG-BRIL-BAG2
    • Protein or peptide: Anhydromuropeptide permease,Soluble cytochrome b562
  • Ligand: DODECYL-BETA-D-MALTOSIDE
  • Ligand: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine
KeywordsMembrane protein / Major Facilitator Family transporter / cell wall / antibiotic resistance
Function / homology
Function and homology information


peptidoglycan transmembrane transporter activity / peptidoglycan turnover / symporter activity / electron transport chain / cell wall organization / periplasmic space / electron transfer activity / iron ion binding / heme binding / plasma membrane
Similarity search - Function
AmpG-like permease/Acetyl-coenzyme A transporter 1 / Major facilitator superfamily / Major Facilitator Superfamily / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562 / MFS transporter superfamily
Similarity search - Domain/homology
Soluble cytochrome b562 / Anhydromuropeptide permease
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.78 Å
AuthorsSverak H / Worrall LJ / Strynadka NCJ
Funding support Canada, 1 items
OrganizationGrant numberCountry
Canadian Institutes of Health Research (CIHR) Canada
CitationJournal: Nat Commun / Year: 2024
Title: Cryo-EM characterization of the anydromuropeptide permease AmpG central to bacterial fitness and β-lactam antibiotic resistance.
Authors: Helena E Sverak / Luke N Yaeger / Liam J Worrall / Condurache M Vacariu / Amy J Glenwright / Marija Vuckovic / Zayni-Dean Al Azawi / Ryan P Lamers / Victoria A Marko / Clarissa Skorupski / ...Authors: Helena E Sverak / Luke N Yaeger / Liam J Worrall / Condurache M Vacariu / Amy J Glenwright / Marija Vuckovic / Zayni-Dean Al Azawi / Ryan P Lamers / Victoria A Marko / Clarissa Skorupski / Arvind S Soni / Martin E Tanner / Lori L Burrows / Natalie Cj Strynadka /
Abstract: Bacteria invest significant resources into the continuous creation and tailoring of their essential protective peptidoglycan (PG) cell wall. Several soluble PG biosynthesis products in the periplasm ...Bacteria invest significant resources into the continuous creation and tailoring of their essential protective peptidoglycan (PG) cell wall. Several soluble PG biosynthesis products in the periplasm are transported to the cytosol for recycling, leading to enhanced bacterial fitness. GlcNAc-1,6-anhydroMurNAc and peptide variants are transported by the essential major facilitator superfamily importer AmpG in Gram-negative pathogens including Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa. Accumulation of GlcNAc-1,6-anhydroMurNAc-pentapeptides also results from β-lactam antibiotic induced cell wall damage. In some species, these products upregulate the β-lactamase AmpC, which hydrolyzes β-lactams to allow for bacterial survival and drug-resistant infections. Here, we have used cryo-electron microscopy and chemical synthesis of substrates in an integrated structural, biochemical, and cellular analysis of AmpG. We show how AmpG accommodates the large GlcNAc-1,6-anhydroMurNAc peptides, including a unique hydrophobic vestibule to the substrate binding cavity, and characterize residues involved in binding that inform the mechanism of proton-mediated transport.
History
DepositionMay 31, 2024-
Header (metadata) releaseDec 18, 2024-
Map releaseDec 18, 2024-
UpdateJun 4, 2025-
Current statusJun 4, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45167.png

Downloads & links

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Map

FileDownload / File: emd_45167.map.gz / Format: CCP4 / Size: 42.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.18 Å/pix.
x 224 pix.
= 264.32 Å		1.18 Å/pix.
x 224 pix.
= 264.32 Å		1.18 Å/pix.
x 224 pix.
= 264.32 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.18 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.3
Minimum - Maximum-1.9803194 - 2.4904635
Average (Standard dev.)0.00061102037 (±0.04093318)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions224224224
Spacing224224224
CellA=B=C: 264.31998 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_45167_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: #2

Fileemd_45167_half_map_2.map
Projections & Slices
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Histogram

Histogram (log scale)

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Sample components

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Entire : AmpG-BRIL-BAG2

EntireName: AmpG-BRIL-BAG2
Components
  • Complex: AmpG-BRIL-BAG2
    • Protein or peptide: Anhydromuropeptide permease,Soluble cytochrome b562
  • Ligand: DODECYL-BETA-D-MALTOSIDE
  • Ligand: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine

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Supramolecule #1: AmpG-BRIL-BAG2

SupramoleculeName: AmpG-BRIL-BAG2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: Complex between AmpG-BRIL and BAG2 antibody
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: Anhydromuropeptide permease,Soluble cytochrome b562

MacromoleculeName: Anhydromuropeptide permease,Soluble cytochrome b562 / type: protein_or_peptide / ID: 1
Details: AmpG with C-terminal BRIL insertion. N-terminal recombinant His-tag cleaved off.
Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 65.32723 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: GSHMSSQYLR IFQQPRSAIL LILGFASGLP LALTSGTLQA WMTVENIDLK TIGFFSLVGQ AYVFKFLWSP LMDRYTPPFF GRRRGWLLA TQILLLVAIA AMGFLEPGTQ LRWMAALAVV IAFCSASQDI VFDAWKTDVL PAEERGAGAA ISVLGYRLGM L VSGGLALW ...String:
GSHMSSQYLR IFQQPRSAIL LILGFASGLP LALTSGTLQA WMTVENIDLK TIGFFSLVGQ AYVFKFLWSP LMDRYTPPFF GRRRGWLLA TQILLLVAIA AMGFLEPGTQ LRWMAALAVV IAFCSASQDI VFDAWKTDVL PAEERGAGAA ISVLGYRLGM L VSGGLALW LADKWLGWQG MYWLMAALLI PCIIATLLAP EPTDTIPVPK TLEQAVVAPL RDFFGRNNAW LILLLIVLYK LG DAFAMSL TTTFLIRGVG FDAGEVGVVN KTLGLLATIV GALYGGILMQ RLSLFRALLI FGILQGASNA GYWLLSITDK HLY SMGAAV FFENLCGGMG TSAFVALLMT LCNKSFSATQ FALLSALSAV GRVYVGPVAG WFVEAHGWST FYLFSVAAAV PGLI LLLVC RQTLEYTRVN DNFISRTAYP AGYAFAMWTL AAGVSLLAVW LLLLTMDALD LTHFSFLPAL LEVGVLVALS GVVLG GLLD YLALRKTHLT ADLEDNWETL NDNLKVIEKA DNAAQVKDAL TKMRAAALDA QKATPPKLED KSPDSPEMKD FRHGFD ILV GQIDDALKLA NEGKVKEAQA AAEQLKTTRN AYIQKYL

UniProtKB: Anhydromuropeptide permease, Soluble cytochrome b562

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Macromolecule #2: DODECYL-BETA-D-MALTOSIDE

MacromoleculeName: DODECYL-BETA-D-MALTOSIDE / type: ligand / ID: 2 / Number of copies: 1 / Formula: LMT
Molecular weightTheoretical: 510.615 Da
Chemical component information

ChemComp-LMT:
DODECYL-BETA-D-MALTOSIDE / detergent*YM

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Macromolecule #3: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine

MacromoleculeName: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine / type: ligand / ID: 3 / Number of copies: 1 / Formula: PEE
Molecular weightTheoretical: 744.034 Da
Chemical component information

ChemComp-PEE:
1,2-dioleoyl-sn-glycero-3-phosphoethanolamine / DOPE, phospholipid*YM

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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: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.78 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 500000
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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