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- EMDB-4470: Spiral structure of E. coli RavA in the RavA-LdcI cage-like complex -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-4470 | |||||||||||||||
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Title | Spiral structure of E. coli RavA in the RavA-LdcI cage-like complex | |||||||||||||||
![]() | Cryo-EM map of Class 2 of the E. coli RavA-LdcI cage-like complex after symmetry expansion and masked 3D classification. | |||||||||||||||
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Function / homology | ![]() lysine decarboxylase / lysine decarboxylase activity / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to catalyse transmembrane movement of substances / amino acid metabolic process / ATP hydrolysis activity / ATP binding / cytoplasm Similarity search - Function | |||||||||||||||
Biological species | ![]() ![]() | |||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 7.8 Å | |||||||||||||||
![]() | Arragain B / Felix J / Malet H / Gutsche I / Jessop M | |||||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural insights into ATP hydrolysis by the MoxR ATPase RavA and the LdcI-RavA cage-like complex. Authors: Matthew Jessop / Benoit Arragain / Roger Miras / Angélique Fraudeau / Karine Huard / Maria Bacia-Verloop / Patrice Catty / Jan Felix / Hélène Malet / Irina Gutsche / ![]() Abstract: The hexameric MoxR AAA+ ATPase RavA and the decameric lysine decarboxylase LdcI form a 3.3 MDa cage, proposed to assist assembly of specific respiratory complexes in E. coli. Here, we show that ...The hexameric MoxR AAA+ ATPase RavA and the decameric lysine decarboxylase LdcI form a 3.3 MDa cage, proposed to assist assembly of specific respiratory complexes in E. coli. Here, we show that inside the LdcI-RavA cage, RavA hexamers adopt an asymmetric spiral conformation in which the nucleotide-free seam is constrained to two opposite orientations. Cryo-EM reconstructions of free RavA reveal two co-existing structural states: an asymmetric spiral, and a flat C2-symmetric closed ring characterised by two nucleotide-free seams. The closed ring RavA state bears close structural similarity to the pseudo two-fold symmetric crystal structure of the AAA+ unfoldase ClpX, suggesting a common ATPase mechanism. Based on these structures, and in light of the current knowledge regarding AAA+ ATPases, we propose different scenarios for the ATP hydrolysis cycle of free RavA and the LdcI-RavA cage-like complex, and extend the comparison to other AAA+ ATPases of clade 7. | |||||||||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 2.9 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 22.3 KB 22.3 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 7.2 KB | Display | ![]() |
Images | ![]() | 261.5 KB | ||
Masks | ![]() | 30.5 MB | ![]() | |
Others | ![]() ![]() | 23.5 MB 23.5 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6q7mMC ![]() 4469C ![]() 6q7lC ![]() 6szaC ![]() 6szbC C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Map
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Annotation | Cryo-EM map of Class 2 of the E. coli RavA-LdcI cage-like complex after symmetry expansion and masked 3D classification. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X: 2.42 Å / Y: 2.42 Å / Z: 2.42001 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
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Projections & Slices |
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Density Histograms |
-Half map: Unfiltered half-map 1.
File | emd_4470_half_map_1.map | ||||||||||||
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Annotation | Unfiltered half-map 1. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Unfiltered half-map 1.
File | emd_4470_half_map_2.map | ||||||||||||
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Annotation | Unfiltered half-map 1. | ||||||||||||
Projections & Slices |
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Density Histograms |
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Sample components
-Entire : Complex of the hexameric MoxR AAA+ ATPase RavA and the decameric ...
Entire | Name: Complex of the hexameric MoxR AAA+ ATPase RavA and the decameric lysine decarboxylase LdcI. |
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Components |
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-Supramolecule #1: Complex of the hexameric MoxR AAA+ ATPase RavA and the decameric ...
Supramolecule | Name: Complex of the hexameric MoxR AAA+ ATPase RavA and the decameric lysine decarboxylase LdcI. type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2 |
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Source (natural) | Organism: ![]() ![]() |
Recombinant expression | Organism: ![]() ![]() |
Molecular weight | Theoretical: 3.3 MDa |
-Macromolecule #1: Inducible lysine decarboxylase
Macromolecule | Name: Inducible lysine decarboxylase / type: protein_or_peptide / ID: 1 / Number of copies: 20 / Enantiomer: LEVO / EC number: lysine decarboxylase |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 81.357008 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MNVIAILNHM GVYFKEEPIR ELHRALERLN FQIVYPNDRD DLLKLIENNA RLCGVIFDWD KYNLELCEEI SKMNENLPLY AFANTYSTL DVSLNDLRLQ ISFFEYALGA AEDIANKIKQ TTDEYINTIL PPLTKALFKY VREGKYTFCT PGHMGGTAFQ K SPVGSLFY ...String: MNVIAILNHM GVYFKEEPIR ELHRALERLN FQIVYPNDRD DLLKLIENNA RLCGVIFDWD KYNLELCEEI SKMNENLPLY AFANTYSTL DVSLNDLRLQ ISFFEYALGA AEDIANKIKQ TTDEYINTIL PPLTKALFKY VREGKYTFCT PGHMGGTAFQ K SPVGSLFY DFFGPNTMKS DISISVSELG SLLDHSGPHK EAEQYIARVF NADRSYMVTN GTSTANKIVG MYSAPAGSTI LI DRNCHKS LTHLMMMSDV TPIYFRPTRN AYGILGGIPQ SEFQHATIAK RVKETPNATW PVHAVITNST YDGLLYNTDF IKK TLDVKS IHFDSAWVPY TNFSPIYEGK CGMSGGRVEG KVIYETQSTH KLLAAFSQAS MIHVKGDVNE ETFNEAYMMH TTTS PHYGI VASTETAAAM MKGNAGKRLI NGSIERAIKF RKEIKRLRTE SDGWFFDVWQ PDHIDTTECW PLRSDSTWHG FKNID NEHM YLDPIKVTLL TPGMEKDGTM SDFGIPASIV AKYLDEHGIV VEKTGPYNLL FLFSIGIDKT KALSLLRALT DFKRAF DLN LRVKNMLPSL YREDPEFYEN MRIQELAQNI HKLIVHHNLP DLMYRAFEVL PTMVMTPYAA FQKELHGMTE EVYLDEM VG RINANMILPY PPGVPLVMPG EMITEESRPV LEFLQMLCEI GAHYPGFETD IHGAYRQADG RYTVKVLKEE SKK |
-Macromolecule #2: ATPase RavA
Macromolecule | Name: ATPase RavA / type: protein_or_peptide / ID: 2 / Number of copies: 6 / Enantiomer: LEVO EC number: Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to catalyse transmembrane movement of substances |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 56.351445 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MAHPHLLAER ISRLSSSLEK GLYERSHAIR LCLLAALSGE SVFLLGPPGI AKSLIARRLK FAFQNARAFE YLMTRFSTPE EVFGPLSIQ ALKDEGRYER LTSGYLPEAE IVFLDEIWKA GPAILNTLLT AINERQFRNG AHVEKIPMRL LVAASNELPE A DSSLEALY ...String: MAHPHLLAER ISRLSSSLEK GLYERSHAIR LCLLAALSGE SVFLLGPPGI AKSLIARRLK FAFQNARAFE YLMTRFSTPE EVFGPLSIQ ALKDEGRYER LTSGYLPEAE IVFLDEIWKA GPAILNTLLT AINERQFRNG AHVEKIPMRL LVAASNELPE A DSSLEALY DRMLIRLWLD KVQDKANFRS MLTSQQDEND NPVPDALQVT DEEYERWQKE IGEITLPDHV FELIFMLRQQ LD KLPDAPY VSDRRWKKAI RLLQASAFFS GRSAVAPVDL ILLKDCLWYD AQSLNLIQQQ IDVLMTGHAW QQQGMLTRLG AIV QRHLQL QQQQSDKTAL TVIRLGGIFS RRQQYQLPVN VTASTLTLLL QKPLKLHDME VVHISFERSA LEQWLSKGGE IRGK LNGIG FAQKLNLEVD SAQHLVVRDV SLQGSTLALP GSSAEGLPGE IKQQLEELES DWRKQHALFS EQQKCLFIPG DWLGR IEAS LQDVGAQIRQ AQQ |
-Macromolecule #3: PYRIDOXAL-5'-PHOSPHATE
Macromolecule | Name: PYRIDOXAL-5'-PHOSPHATE / type: ligand / ID: 3 / Number of copies: 20 / Formula: PLP |
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Molecular weight | Theoretical: 247.142 Da |
Chemical component information | ![]() ChemComp-PLP: |
-Macromolecule #4: ADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 4 / Number of copies: 5 / Formula: ADP |
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Molecular weight | Theoretical: 427.201 Da |
Chemical component information | ![]() ChemComp-ADP: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.98 mg/mL |
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Buffer | pH: 7.9 Details: 20 mM Tris pH 7.9, 300 mM NaCl, 2 mM ADP, 10 mM MgCl 2 , 0.1 mM PLP and 1 mM DTT |
Vitrification | Cryogen name: ETHANE |
Details | Concentrations of LdcI and RavA were 0.38 mg/ml (4.67 microM) and 0.6 mg/ml (10.64 microM) respectively. |
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Electron microscopy
Microscope | FEI POLARA 300 |
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Details | Data collection was performed on an FEI Polara microscope operated at 300 kV. Movies of 40 frames were collected with a total exposure time of 8s and a total dose of 40e-/Angstrom^2 on a K2 summit direct electron detector (Gatan) at a magnification of 41270x, corresponding to 1.21 Angstrom/pixel at the specimen level. |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 3-40 / Number real images: 1819 / Average exposure time: 8.0 sec. / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated magnification: 41270 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | ![]() Model: Tecnai Polara / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Initial model | (PDB ID: , ) |
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Details | Local resolution estimation and subsequent filtering of maps were performed in Relion3.0. For fitting of atomic models in the resulting filtered maps, we used the previously-determined X-ray structures of LdcI (PDB ID: 3N75) (Kanjee et al., 2011) and RavA (PDB ID: 3NBX) (El Bakkouri et al., 2010). In each map, two decameric LdcI molecules extracted from PDB 3N75 and one spiral RavA hexamer extracted from a continuous RavA helix generated from PDB 3NBX were fitted separately using iMODFIT (Lopez-Blanco and Chacon, 2013), followed by a single round of B-factor (ADP) refinement in Phenix. |
Refinement | Protocol: FLEXIBLE FIT |
Output model | ![]() PDB-6q7m: |