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- EMDB-34002: Spiral pentamer of the substrate-free Lon protease with a S678A m... -
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Open data
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Basic information
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Title | Spiral pentamer of the substrate-free Lon protease with a S678A mutation | |||||||||
![]() | Pentameric state of MtaLon-S678A with ADP | |||||||||
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Function / homology | ![]() ![]() ![]() ![]() ![]() Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | ![]() ![]() | |||||||||
![]() | Li S / Hsieh KY / Kuo CI / Lee SH / Ho MR / Wang CH / Zhang K / Chang CI | |||||||||
Funding support | ![]()
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![]() | ![]() Title: A 5+1 assemble-to-activate mechanism of the Lon proteolytic machine. Authors: Shanshan Li / Kan-Yen Hsieh / Chiao-I Kuo / Tzu-Chi Lin / Szu-Hui Lee / Yi-Ru Chen / Chun-Hsiung Wang / Meng-Ru Ho / See-Yeun Ting / Kaiming Zhang / Chung-I Chang / ![]() ![]() Abstract: Many AAA+ (ATPases associated with diverse cellular activities) proteins function as protein or DNA remodelers by threading the substrate through the central pore of their hexameric assemblies. In ...Many AAA+ (ATPases associated with diverse cellular activities) proteins function as protein or DNA remodelers by threading the substrate through the central pore of their hexameric assemblies. In this ATP-dependent translocating state, the substrate is gripped by the pore loops of the ATPase domains arranged in a universal right-handed spiral staircase organization. However, the process by which a AAA+ protein is activated to adopt this substrate-pore-loop arrangement remains unknown. We show here, using cryo-electron microscopy (cryo-EM), that the activation process of the Lon AAA+ protease may involve a pentameric assembly and a substrate-dependent incorporation of the sixth protomer to form the substrate-pore-loop contacts seen in the translocating state. Based on the structural results, we design truncated monomeric mutants that inhibit Lon activity by binding to the native pentamer and demonstrated that expressing these monomeric mutants in Escherichia coli cells containing functional Lon elicits specific phenotypes associated with lon deficiency, including the inhibition of persister cell formation. These findings uncover a substrate-dependent assembly process for the activation of a AAA+ protein and demonstrate a targeted approach to selectively inhibit its function within cells. | |||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 62.5 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 18.4 KB 18.4 KB | Display Display | ![]() |
Images | ![]() | 129.9 KB | ||
Filedesc metadata | ![]() | 6.4 KB | ||
Others | ![]() ![]() | 116 MB 116 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 7ypjMC ![]() 7yphC ![]() 7ypiC ![]() 7ypkC ![]() 7yuhC ![]() 7yumC ![]() 7yupC ![]() 7yutC ![]() 7yuuC ![]() 7yuvC ![]() 7yuwC ![]() 7yuxC ![]() 8k3yC M: atomic model generated by this map C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Pentameric state of MtaLon-S678A with ADP | ||||||||||||||||||||
Voxel size | X=Y=Z: 1.061 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: Half map A
File | emd_34002_half_map_1.map | ||||||||||||
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Annotation | Half map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map B
File | emd_34002_half_map_2.map | ||||||||||||
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Annotation | Half map B | ||||||||||||
Projections & Slices |
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Density Histograms |
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Sample components
-Entire : Substrate-free pentamer of MtaLon-S678A
Entire | Name: Substrate-free pentamer of MtaLon-S678A |
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Components |
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-Supramolecule #1: Substrate-free pentamer of MtaLon-S678A
Supramolecule | Name: Substrate-free pentamer of MtaLon-S678A / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 440 KDa |
-Macromolecule #1: Lon protease
Macromolecule | Name: Lon protease / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO / EC number: ![]() |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 88.538594 KDa |
Recombinant expression | Organism: ![]() ![]() ![]() |
Sequence | String: MRLELPVIPL RNTVILPHTT TPVDVGRAKS KRAVEEAMGA DRLIFLVAQR DPEVDDPAPD DLYTWGVQAV VKQAMRLPDG TLQVMVEAR ARAQVTDYIP GPYLRARGEV FSEIFPIDEA VVRVLVEELK EAFEKYVANH KSLRLDRYQL EAVKGTSDPA M LADTIAYH ...String: MRLELPVIPL RNTVILPHTT TPVDVGRAKS KRAVEEAMGA DRLIFLVAQR DPEVDDPAPD DLYTWGVQAV VKQAMRLPDG TLQVMVEAR ARAQVTDYIP GPYLRARGEV FSEIFPIDEA VVRVLVEELK EAFEKYVANH KSLRLDRYQL EAVKGTSDPA M LADTIAYH ATWTVAEKQE ILELTDLEAR LKKVLGLLSR DLERFELDKR VAQRVKEQMD TNQREYYLRE QMKAIQKELG GE DGLSDLE ALRKKIEEVG MPEAVKTKAL KELDRLERMQ QGSPEATVAR TYLDWLTEVP WSKADPEVLD INHTRQVLDE DHY GLKDVK ERILEYLAVR QLTQGLDVRN KAPILVLVGP PGVGKTSLGR SIARSMNRKF HRISLGGVRD EAEIRGHRRT YIGA MPGKL IHAMKQVGVI NPVILLDEID KMSSDWRGDP ASAMLEVLDP EQNNTFTDHY LDVPYDLSKV FFITTANTLQ TIPRP LLDR MEVIEIPGYT NMEKQAIARQ YLWPKQVRES GMEGRIEVTD AAILRVISEY TREAGVRGLE RELGKIARKG AKFWLE GAW EGLRTIDASD IPTYLGIPRY RPDKAETEPQ VGTAQGLAWT PVGGTLLTIE VAAVPGSGKL SLTGQLGEVM KESAQAA LT YLRAHTQDYG LPEDFYNKVD LHVHVPDGAT PKDGPAAGIT MATAIASALS RRPARMDIAM TGEVSLRGKV MPIGGVKE K LLAAHQAGIH KIVLPKDNEA QLEELPKEVL EGLEIKLVED VGEVLEYLLL PEPTMPPVVQ PSDNRQQPGA GA UniProtKB: ![]() |
-Macromolecule #2: ADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 2 / Number of copies: 4 / Formula: ADP |
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Molecular weight | Theoretical: 427.201 Da |
Chemical component information | ![]() ChemComp-ADP: |
-Experimental details
-Structure determination
Method | ![]() |
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Aggregation state | particle |
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Sample preparation
Concentration | 0.5 mg/mL | ||||||||||||||||||
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Buffer | pH: 8 Component:
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Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. | ||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K | ||||||||||||||||||
Details | alpha-S1-casein was incubated with Lon protease |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD![]() |
Sample stage | Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 2 / Number real images: 12546 / Average exposure time: 2.52 sec. / Average electron dose: 59.0 e/Å2 |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
Particle selection | Number selected: 1756270 |
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Startup model | Type of model: OTHER |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.2) |
Final angle assignment | Type: PROJECTION MATCHING / Software - Name: cryoSPARC (ver. 3.2) |
Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.2) / Number images used: 486028 |