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- EMDB-29045: Klebsiella pneumoniae AcrB multidrug efflux pump apo form -

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Basic information

Entry
Database: EMDB / ID: EMD-29045
TitleKlebsiella pneumoniae AcrB multidrug efflux pump apo form
Map data
Sample
  • Complex: AcrBAcriflavine resistance protein family
    • Protein or peptide: Efflux pump membrane transporter
KeywordsKlebsiella pneumoniae / AcrB multidrug efflux pump / MEMBRANE PROTEIN
Function / homologyHydrophobe/amphiphile efflux-1 HAE1 / Acriflavin resistance protein / Multidrug efflux transporter AcrB TolC docking domain, DN/DC subdomains / AcrB/AcrD/AcrF family / xenobiotic transport / efflux transmembrane transporter activity / plasma membrane / Efflux pump membrane transporter
Function and homology information
Biological speciesKlebsiella pneumoniae (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.82 Å
AuthorsZhang Z
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01AI145069 United States
CitationJournal: mBio / Year: 2023
Title: Cryo-EM Structures of the Klebsiella pneumoniae AcrB Multidrug Efflux Pump.
Authors: Zhemin Zhang / Christopher E Morgan / Robert A Bonomo / Edward W Yu /
Abstract: The continued challenges of the COVID-19 pandemic combined with the growing problem of antimicrobial-resistant bacterial infections has severely impacted global health. Specifically, the Gram- ...The continued challenges of the COVID-19 pandemic combined with the growing problem of antimicrobial-resistant bacterial infections has severely impacted global health. Specifically, the Gram-negative pathogen Klebsiella pneumoniae is one of the most prevalent causes of secondary bacterial infection in COVID-19 patients, with approximately an 83% mortality rate observed among COVID-19 patients with these bacterial coinfections. K. pneumoniae belongs to the ESKAPE group of pathogens, a group that commonly gives rise to severe infections that are often life-threatening. Recently, K. pneumoniae carbapenemase (KPC)-producing K. pneumoniae has drawn wide public attention, as the mortality rate for this infection can be as high as 71%. The most predominant and clinically important multidrug efflux system in K. pneumoniae is the acriflavine resistance B (AcrB) multidrug efflux pump. This pump mediates resistance to different classes of structurally diverse antimicrobial agents, including quinolones, β-lactams, tetracyclines, macrolides, aminoglycosides, and chloramphenicol. We here report single-particle cryo-electron microscopy (cryo-EM) structures of K. pneumoniae AcrB, in both the absence and the presence of the antibiotic erythromycin. These structures allow us to elucidate specific pump-drug interactions and pinpoint exactly how this pump recognizes antibiotics. Klebsiella pneumoniae has emerged as one of the most problematic and highly antibiotic-resistant pathogens worldwide. It is the second most common causative agent involved in secondary bacterial infection in COVID-19 patients. K. pneumoniae carbapenemase (KPC)-producing K. pneumoniae is a major concern in global public health because of the high mortality rate of this infection. Its drug resistance is due, in a significant part, to active efflux of these bactericides, a major mechanism that K. pneumoniae uses to resist to the action of multiple classes of antibiotics. Here, we report cryo-electron microscopy (cryo-EM) structures of the prevalent and clinically important K. pneumoniae AcrB multidrug efflux pump, in both the absence and the presence of the erythromycin antibiotic. These structures allow us to understand the action mechanism for drug recognition in this pump. Our studies will ultimately inform an era in structure-guided drug design to combat multidrug resistance in these Gram-negative pathogens.
History
DepositionDec 8, 2022-
Header (metadata) releaseApr 5, 2023-
Map releaseApr 5, 2023-
UpdateJul 12, 2023-
Current statusJul 12, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_29045.png

Downloads & links

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Map

FileDownload / File: emd_29045.map.gz / Format: CCP4 / Size: 166.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.07 Å
Density
Contour LevelBy AUTHOR: 0.2
Minimum - Maximum-0.8715951 - 1.7700173
Average (Standard dev.)-0.00027846548 (±0.05068929)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions352352352
Spacing352352352
CellA=B=C: 376.64 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_29045_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_29045_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : AcrB

EntireName: AcrBAcriflavine resistance protein family
Components
  • Complex: AcrBAcriflavine resistance protein family
    • Protein or peptide: Efflux pump membrane transporter

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Supramolecule #1: AcrB

SupramoleculeName: AcrB / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Klebsiella pneumoniae (bacteria)

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Macromolecule #1: Efflux pump membrane transporter

MacromoleculeName: Efflux pump membrane transporter / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Klebsiella pneumoniae (bacteria)
Molecular weightTheoretical: 113.404805 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MPNFFIDRPI FAWVIAIIIM LAGGLSILKL PVAQYPTIAP PAISITAMYP GADAETVQNT VTQVIEQNMN GIDHLMYMSS NGDSTGTAT ITLTFESGTD PDIAQVQVQN KLALATPLLP QEVQQQGISV EKASSSFLMV VGVINTNGTM NQDDISDYVA A NMKDPISR ...String:
MPNFFIDRPI FAWVIAIIIM LAGGLSILKL PVAQYPTIAP PAISITAMYP GADAETVQNT VTQVIEQNMN GIDHLMYMSS NGDSTGTAT ITLTFESGTD PDIAQVQVQN KLALATPLLP QEVQQQGISV EKASSSFLMV VGVINTNGTM NQDDISDYVA A NMKDPISR TSGVGDVQLF GSQYAMRIWM DPNKLNNFQL TPVDVISALK AQNAQVAAGQ LGGTPPVKGQ QLNASIIAQT RL TNTEEFG NILLKVNQDG SQVRLRDVAK IELGGESYDV VAKFNGQPAS GLGIKLATGA NALDTANAIR AELAKMEPFF PSG MKIVYP YDTTPFVKIS IHEVVKTLVE AIILVFLVMY LFLQNFRATL IPTIAVPVVL LGTFAVLAAF GFSINTLTMF GMVL AIGLL VDDAIVVVEN VERVMAEEGL PPKEATRKSM GQIQGALVGI AMVLSAVFIP MAFFGGSTGA IYRQFSITIV SAMAL SVLV ALILTPALCA TMLKPIQKGS HGATTGFFGW FNRMFDKSTH HYTDSVGNIL RSTGRYLVLY LIIVVGMAWL FVRLPS SFL PDEDQGVFLS MAQLPAGATQ ERTQKVLDEM TNYYLTKEKD NVESVFAVNG FGFAGRGQNT GIAFVSLKDW SQRPGEE NK VEAITARAMG YFSQIKDAMV FAFNLPAIVE LGTATGFDFE LIDQGGLGHE KLTQARNQLF GMVAQHPDVL TGVRPNGL E DTPQFKIDID QEKAQALGVS ISDINTTLGA AWGGSYVNDF IDRGRVKKVY IMSEAKYRML PEDIGKWYVR GSDGQMVPF SAFSTSRWEY GSPRLERYNG LPSLEILGQA APGKSTGEAM ALMEELAGKL PSGIGYDWTG MSYQERLSGN QAPALYAISL IVVFLCLAA LYESWSIPFS VMLVVPLGVV GALLAATFRG LTNDVYFQVG LLTTIGLSAK NAILIVEFAK DLMEKEGKGL I EATLEAVR MRLRPILMTS LAFILGVMPL VISSGAGSGA QNAVGTGVMG GMVTATILAI FFVPVFFVVV RRRFSKKSED IE HSHQVEH H

UniProtKB: Efflux pump membrane transporter

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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
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 81000
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 35.5 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.82 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 64539
FSC plot (resolution estimation)

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