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- EMDB-12533: Cryo-EM structure of the cytochrome bd oxidase from M. tuberculos... -

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Basic information

Entry
Database: EMDB / ID: EMD-12533
TitleCryo-EM structure of the cytochrome bd oxidase from M. tuberculosis in presence of Aurachin D at 3.3 A resolution
Map dataPostprocess map Applied b-factor: -127
Sample
  • Complex: Cryo-EM structure of the cytochrome bd oxidase from M. tuberculosis in presence of Aurachin D at 3.3 A resolution
    • Protein or peptide: Probable integral membrane cytochrome D ubiquinol oxidase (Subunit II) CydB (Cytochrome BD-I oxidase subunit II)
    • Protein or peptide: Probable integral membrane cytochrome D ubiquinol oxidase (Subunit I) CydA (Cytochrome BD-I oxidase subunit I)
Function / homology
Function and homology information


cytochrome complex / aerobic electron transport chain / oxidoreductase activity, acting on diphenols and related substances as donors, oxygen as acceptor / electron transfer activity / membrane => GO:0016020 / heme binding / metal ion binding / plasma membrane
Similarity search - Function
Cytochrome ubiquinol oxidase subunit 1 / Cytochrome ubiquinol oxidase subunit 2 / Cytochrome bd terminal oxidase subunit I / Cytochrome bd terminal oxidase subunit II
Similarity search - Domain/homology
Probable integral membrane cytochrome D ubiquinol oxidase (Subunit I) CydA (Cytochrome BD-I oxidase subunit I) / Probable integral membrane cytochrome D ubiquinol oxidase (Subunit II) CydB (Cytochrome BD-I oxidase subunit II)
Similarity search - Component
Biological speciesMycobacterium tuberculosis H37Rv (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsSafarian S / Wu D / Krause KL / Michel H
Funding support Germany, New Zealand, 4 items
OrganizationGrant numberCountry
German Research Foundation (DFG) Germany
Max Planck SocietyNobel Laureate Fellowship Germany
Marsden Fund New Zealand
Royal Society of New ZealandCatalyst grant New Zealand
CitationJournal: Nat Commun / Year: 2021
Title: The cryo-EM structure of the bd oxidase from M. tuberculosis reveals a unique structural framework and enables rational drug design to combat TB.
Authors: Schara Safarian / Helen K Opel-Reading / Di Wu / Ahmad R Mehdipour / Kiel Hards / Liam K Harold / Melanie Radloff / Ian Stewart / Sonja Welsch / Gerhard Hummer / Gregory M Cook / Kurt L ...Authors: Schara Safarian / Helen K Opel-Reading / Di Wu / Ahmad R Mehdipour / Kiel Hards / Liam K Harold / Melanie Radloff / Ian Stewart / Sonja Welsch / Gerhard Hummer / Gregory M Cook / Kurt L Krause / Hartmut Michel /
Abstract: New drugs are urgently needed to combat the global TB epidemic. Targeting simultaneously multiple respiratory enzyme complexes of Mycobacterium tuberculosis is regarded as one of the most effective ...New drugs are urgently needed to combat the global TB epidemic. Targeting simultaneously multiple respiratory enzyme complexes of Mycobacterium tuberculosis is regarded as one of the most effective treatment options to shorten drug administration regimes, and reduce the opportunity for the emergence of drug resistance. During infection and proliferation, the cytochrome bd oxidase plays a crucial role for mycobacterial pathophysiology by maintaining aerobic respiration at limited oxygen concentrations. Here, we present the cryo-EM structure of the cytochrome bd oxidase from M. tuberculosis at 2.5 Å. In conjunction with atomistic molecular dynamics (MD) simulation studies we discovered a previously unknown MK-9-binding site, as well as a unique disulfide bond within the Q-loop domain that defines an inactive conformation of the canonical quinol oxidation site in Actinobacteria. Our detailed insights into the long-sought atomic framework of the cytochrome bd oxidase from M. tuberculosis will form the basis for the design of highly specific drugs to act on this enzyme.
History
DepositionMar 3, 2021-
Header (metadata) releaseSep 22, 2021-
Map releaseSep 22, 2021-
UpdateSep 22, 2021-
Current statusSep 22, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0196
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.0196
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_12533.png

Downloads & links

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Map

FileDownload / File: emd_12533.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationPostprocess map Applied b-factor: -127
Voxel sizeX=Y=Z: 0.837 Å
Density
Contour LevelBy AUTHOR: 0.0196 / Movie #1: 0.0196
Minimum - Maximum-0.06445777 - 0.10830398
Average (Standard dev.)7.058119e-05 (±0.0022009332)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 214.272 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.8370.8370.837
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z214.272214.272214.272
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.0640.1080.000

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Supplemental data

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Half map: #1

Fileemd_12533_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_12533_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Cryo-EM structure of the cytochrome bd oxidase from M. tuberculos...

EntireName: Cryo-EM structure of the cytochrome bd oxidase from M. tuberculosis in presence of Aurachin D at 3.3 A resolution
Components
  • Complex: Cryo-EM structure of the cytochrome bd oxidase from M. tuberculosis in presence of Aurachin D at 3.3 A resolution
    • Protein or peptide: Probable integral membrane cytochrome D ubiquinol oxidase (Subunit II) CydB (Cytochrome BD-I oxidase subunit II)
    • Protein or peptide: Probable integral membrane cytochrome D ubiquinol oxidase (Subunit I) CydA (Cytochrome BD-I oxidase subunit I)

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Supramolecule #1: Cryo-EM structure of the cytochrome bd oxidase from M. tuberculos...

SupramoleculeName: Cryo-EM structure of the cytochrome bd oxidase from M. tuberculosis in presence of Aurachin D at 3.3 A resolution
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria)
Recombinant expressionOrganism: Mycolicibacterium smegmatis MC2 155 (bacteria) / Recombinant plasmid: pYUB28b
Molecular weightExperimental: 91.5 KDa

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Macromolecule #1: Probable integral membrane cytochrome D ubiquinol oxidase (Subuni...

MacromoleculeName: Probable integral membrane cytochrome D ubiquinol oxidase (Subunit II) CydB (Cytochrome BD-I oxidase subunit II)
type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria)
Recombinant expressionOrganism: Mycolicibacterium smegmatis MC2 155 (bacteria)
SequenceString: MVLQELWFGV IAALFLGFFI LEGFDFGVGM LMAPFAHVGM GDPETHRRTA LNTIGPVWDG NEVWLITAGA AIFAAFPGWY ATVFSALYLP LLAILFGMIL RAVAIEWRGK IDDPKWRTGA DFGIAAGSWL PALLWGVAFA ILVRGLPVDA NGHVALSIPD VLNAYTLLGG ...String:
MVLQELWFGV IAALFLGFFI LEGFDFGVGM LMAPFAHVGM GDPETHRRTA LNTIGPVWDG NEVWLITAGA AIFAAFPGWY ATVFSALYLP LLAILFGMIL RAVAIEWRGK IDDPKWRTGA DFGIAAGSWL PALLWGVAFA ILVRGLPVDA NGHVALSIPD VLNAYTLLGG LATAGLFSLY GAVFIALKTS GPIRDDAYRF AVWLSLPVAG LVAGFGLWTQ LAYGKDWTWL VLAVAGCAQA AATVLVWRRV SDGWAFMCTL IVVAAVVVLL FGALYPNLVP STLNPQWSLT IHNASSTPYT LKIMTWVTAF FAPLTVAYQT WTYWVFRQRI SAERIPPPTG LARRAP

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Macromolecule #2: Probable integral membrane cytochrome D ubiquinol oxidase (Subuni...

MacromoleculeName: Probable integral membrane cytochrome D ubiquinol oxidase (Subunit I) CydA (Cytochrome BD-I oxidase subunit I)
type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium tuberculosis H37Rv (bacteria)
Recombinant expressionOrganism: Mycolicibacterium smegmatis MC2 155 (bacteria)
SequenceString: MNVVDISRWQ FGITTVYHFI FVPLTIGLAP LIAVMQTLWV VTDNPAWYRL TKFFGKLFLI NFAIGVATGI VQEFQFGMNW SEYSRFVGDV FGAPLAMEGL AAFFFESTFI GLWIFGWNRL PRLVHLACIW IVAIAVNVSA FFIIAANSFM QHPVGAHYNP TTGRAELSSI ...String:
MNVVDISRWQ FGITTVYHFI FVPLTIGLAP LIAVMQTLWV VTDNPAWYRL TKFFGKLFLI NFAIGVATGI VQEFQFGMNW SEYSRFVGDV FGAPLAMEGL AAFFFESTFI GLWIFGWNRL PRLVHLACIW IVAIAVNVSA FFIIAANSFM QHPVGAHYNP TTGRAELSSI VVLLTNNTAQ AAFTHTVSGA LLTAGTFVAA VSAWWLVRSS TTHADSDTQA MYRPATILGC WVALAATAGL LFTGDHQGKL MFQQQPMKMA SAESLCDTQT DPNFSVLTVG RQNNCDSLTR VIEVPYVLPF LAEGRISGVT LQGIRDLQQE YQQRFGPNDY RPNLFVTYWS FRMMIGLMAI PVLFALIALW LTRGGQIPNQ RWFSWLALLT MPAPFLANSA GWVFTEMGRQ PWVVVPNPTG DQLVRLTVKA GVSDHSATVV ATSLLMFTLV YAVLAVIWCW LLKRYIVEGP LEHDAEPAAH GAPRDDEVAP LSFAY

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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

Concentration1.5 mg/mL
BufferpH: 7
Component:
ConcentrationNameFormula
20.0 mMHepes
100.0 mMNaClSodium chloride
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Details: 15 mA
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV / Details: Blot force 20.
DetailsMonodisperse sample of cytochrome bd oxidase reconstituted in lipid nanodiscs 1D1

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 96000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number real images: 7401 / Average exposure time: 5.0 sec. / Average electron dose: 15.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 763298
CTF correctionSoftware - Name: CTFFIND
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final 3D classificationNumber classes: 4 / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final reconstructionNumber classes used: 1 / Algorithm: EXACT BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 194987
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL / Overall B value: 94 / Target criteria: Correlation coefficient

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