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- EMDB-15243: Mycobacterium tuberculosis ClpC1 hexamer structure bound to the n... -

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Basic information

Entry
Database: EMDB / ID: EMD-15243
TitleMycobacterium tuberculosis ClpC1 hexamer structure bound to the natural product antibiotic ecumicin (class 2)
Map dataMycobacterium tuberculosis ClpC1 with added Ecumicin, Class 2
Sample
  • Complex: Mycobacterium tuberculosis ClpC1 bound to the natural product antibiotic Ecumicin
    • Protein or peptide: Mycobacterium tuberculosis ClpC1 with added Ecumicin, Class 2
Keywordshexamer / tuberculosis / drug target / protein quality control / CHAPERONE
Biological speciesMycobacterium tuberculosis (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 8.59 Å
AuthorsFelix J / Fraga H / Gragera M / Bueno T / Weinhaeupl K
Funding supportEuropean Union, 3 items
OrganizationGrant numberCountry
Other governmentiNEXT Discovery (871037)European Union
Other governmentCRIOMECORR project (ESFRI-2019-01-CSIC-16)European Union
Other governmentInstruct-ERIC (PID 15689)European Union
CitationJournal: J Biol Chem / Year: 2022
Title: Structure of the drug target ClpC1 unfoldase in action provides insights on antibiotic mechanism of action.
Authors: Katharina Weinhäupl / Marcos Gragera / M Teresa Bueno-Carrasco / Rocío Arranz / Olga Krandor / Tatos Akopian / Raquel Soares / Eric Rubin / Jan Felix / Hugo Fraga /
Abstract: The unfoldase ClpC1 is one of the most exciting drug targets against tuberculosis. This AAA+ unfoldase works in cooperation with the ClpP1P2 protease and is the target of at least four natural ...The unfoldase ClpC1 is one of the most exciting drug targets against tuberculosis. This AAA+ unfoldase works in cooperation with the ClpP1P2 protease and is the target of at least four natural product antibiotics: cyclomarin, ecumicin, lassomycin, and rufomycin. Although these molecules are promising starting points for drug development, their mechanisms of action remain largely unknown. Taking advantage of a middle domain mutant, we determined the first structure of Mycobacterium tuberculosis ClpC1 in its apo, cyclomarin-, and ecumicin-bound states via cryo-EM. The obtained structure displays features observed in other members of the AAA+ family and provides a map for further drug development. While the apo and cyclomarin-bound structures are indistinguishable and have N-terminal domains that are invisible in their respective EM maps, around half of the ecumicin-bound ClpC1 particles display three of their six N-terminal domains in an extended conformation. Our structural observations suggest a mechanism where ecumicin functions by mimicking substrate binding, leading to ATPase activation and changes in protein degradation profile.
History
DepositionJun 24, 2022-
Header (metadata) releaseMay 31, 2023-
Map releaseMay 31, 2023-
UpdateMay 31, 2023-
Current statusMay 31, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15243.png

Downloads & links

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Map

FileDownload / File: emd_15243.map.gz / Format: CCP4 / Size: 59.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMycobacterium tuberculosis ClpC1 with added Ecumicin, Class 2
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.71 Å/pix.
x 250 pix.
= 427.5 Å		1.71 Å/pix.
x 250 pix.
= 427.5 Å		1.71 Å/pix.
x 250 pix.
= 427.5 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.71 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.55
Minimum - Maximum-1.1318382 - 1.8125468
Average (Standard dev.)0.0014867745 (±0.08275968)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions250250250
Spacing250250250
CellA=B=C: 427.5 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_15243_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_15243_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Mycobacterium tuberculosis ClpC1 bound to the natural product ant...

EntireName: Mycobacterium tuberculosis ClpC1 bound to the natural product antibiotic Ecumicin
Components
  • Complex: Mycobacterium tuberculosis ClpC1 bound to the natural product antibiotic Ecumicin
    • Protein or peptide: Mycobacterium tuberculosis ClpC1 with added Ecumicin, Class 2

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Supramolecule #1: Mycobacterium tuberculosis ClpC1 bound to the natural product ant...

SupramoleculeName: Mycobacterium tuberculosis ClpC1 bound to the natural product antibiotic Ecumicin
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Mycobacterium tuberculosis (bacteria)
Molecular weightTheoretical: 567 KDa

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Macromolecule #1: Mycobacterium tuberculosis ClpC1 with added Ecumicin, Class 2

MacromoleculeName: Mycobacterium tuberculosis ClpC1 with added Ecumicin, Class 2
type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Mycobacterium tuberculosis (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MFERFTDRAR RVVVLAQEEA RMLNHNYIGT EHILLGLIHE GEGVAAKSLE SLGISLEGVR SQVEEIIGQ GQQAPSGHIP FTPRAKKVLE LSLREALQLG HNYIGTEHIL LGLIREGEGV A AQVLVKLG AELTRVRQQV IQLLSGYQGK EAAEAGTGGR GGESGSPSTS ...String:
MFERFTDRAR RVVVLAQEEA RMLNHNYIGT EHILLGLIHE GEGVAAKSLE SLGISLEGVR SQVEEIIGQ GQQAPSGHIP FTPRAKKVLE LSLREALQLG HNYIGTEHIL LGLIREGEGV A AQVLVKLG AELTRVRQQV IQLLSGYQGK EAAEAGTGGR GGESGSPSTS LVLDQFGRNL TA AAMEGKL DPVIGREKEI ERVMQVLSRR TKNNPVLIGE PGVGKTAVVE GLAQAIVHGE VPE TLKDKQ LYTLDLGSLV AGSRYRGDFE ERLKKVLKEI NTRGDIILFI DALHTLVGAG AAEG AIDAA SILKPKLARG ELQTIGATTL DEYRKYIEKD AALERRFQPV QVGEPTVEHT IEILK GLRD RYEAHHRVSI TDAAMVAAAT LADRYINDRF LPDKAIDLID EAGARMRIRR MTAPPD LRE FDEKIAEARR EKESAIDAQD FEKAASLRDR EKTLVAQRAE REKQWRSGDL DVVAEVD DE QIAEVLGNWT GIPVFKLTEA ETTRLLRMEE ELHKRIIGQE DAVKAVSKAI RRTRAGLK D PKRPSGSFIF AGPSGVGKTE LSKALANFLF GDDDALIQID MGEFHDRFTA SRLFGAPPG YVGYEEGGQL TEKVRRKPFS VVLFDAIEKA HQEIYNSLLQ VLEDGRLTDG QGRTVDFKNT VLIFTSNLG TSDISKPVGL GFSKGGGEND YERMKQKVND ELKKHFRPEF LNRIDDIIVF H QLTREEII RMVDLMISRV AGQLKSKDMA LVLTDAAKAL LAKRGFDPVL GARPLRRTIQ RE IEDQLSE KILFEEVGPG QVVTVDVDNW DGEGPGEDAV FTFTGTRKPP AEPDLAKAGA HSA GGPEPA ARLEHHHHHH

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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 mg/mL
BufferpH: 7.4
Details: Hepes pH 7.4 50 mM, NaCl 100 mM, 10 mM MgCl2, ATP 1mM
VitrificationCryogen name: ETHANE
Details10 microM ClpC1 with 30 microM Ecumicin

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Average exposure time: 40.0 sec. / Average electron dose: 32.2 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: OTHER / Nominal defocus max: 3.1 µm / Nominal defocus min: 1.2 µm
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Resolution.type: BY AUTHOR / Resolution: 8.59 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. v3.3.2) / Number images used: 30976
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v3.3.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v3.3.2)
Final 3D classificationSoftware - Name: cryoSPARC (ver. v3.3.2)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

7abr

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