[English] 日本語
Yorodumi- EMDB-6303: The cryo-EM structure of Meiothermus taiwanensis Lon protease wit... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-6303 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | The cryo-EM structure of Meiothermus taiwanensis Lon protease with Mg2+ | |||||||||
Map data | Reconstruction of Meiothermus taiwanensis LonA protease | |||||||||
Sample |
| |||||||||
Keywords | ATP-dependent protease / sequestered degradation chamber / AAA+ domain / protease domain | |||||||||
Biological species | Meiothermus taiwanensis (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / negative staining / Resolution: 12.6 Å | |||||||||
Authors | Su SC / Chang YC / Chang CI | |||||||||
Citation | Journal: Structure / Year: 2016 Title: Structural Basis for the Magnesium-Dependent Activation and Hexamerization of the Lon AAA+ Protease. Authors: Shih-Chieh Su / Chien-Chu Lin / Hui-Chung Tai / Mu-Yueh Chang / Meng-Ru Ho / C Satheesan Babu / Jiahn-Haur Liao / Shih-Hsiung Wu / Yuan-Chih Chang / Carmay Lim / Chung-I Chang / Abstract: The Lon AAA+ protease (LonA) plays important roles in protein homeostasis and regulation of diverse biological processes. LonA behaves as a homomeric hexamer in the presence of magnesium (Mg(2+)) and ...The Lon AAA+ protease (LonA) plays important roles in protein homeostasis and regulation of diverse biological processes. LonA behaves as a homomeric hexamer in the presence of magnesium (Mg(2+)) and performs ATP-dependent proteolysis. However, it is also found that LonA can carry out Mg(2+)-dependent degradation of unfolded protein substrate in an ATP-independent manner. Here we show that in the presence of Mg(2+) LonA forms a non-secluded hexameric barrel with prominent openings, which explains why Mg(2+)-activated LonA can operate as a diffusion-based chambered protease to degrade unstructured protein and peptide substrates efficiently in the absence of ATP. A 1.85 Å crystal structure of Mg(2+)-activated protease domain reveals Mg(2+)-dependent remodeling of a substrate-binding loop and a potential metal-binding site near the Ser-Lys catalytic dyad, supported by biophysical binding assays and molecular dynamics simulations. Together, these findings reveal the specific roles of Mg(2+) in the molecular assembly and activation of LonA. | |||||||||
History |
|
-Structure visualization
Movie |
Movie viewer |
---|---|
Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_6303.map.gz | 6.8 MB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-6303-v30.xml emd-6303.xml | 10 KB 10 KB | Display Display | EMDB header |
Images | emd_6303.tiff | 240 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-6303 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-6303 | HTTPS FTP |
-Validation report
Summary document | emd_6303_validation.pdf.gz | 78.5 KB | Display | EMDB validaton report |
---|---|---|---|---|
Full document | emd_6303_full_validation.pdf.gz | 77.7 KB | Display | |
Data in XML | emd_6303_validation.xml.gz | 493 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6303 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-6303 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|
-Map
File | Download / File: emd_6303.map.gz / Format: CCP4 / Size: 7.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Reconstruction of Meiothermus taiwanensis LonA protease | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.65 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
|
-Supplemental data
-Sample components
-Entire : Meiothermus taiwanensis LonA protease
Entire | Name: Meiothermus taiwanensis LonA protease |
---|---|
Components |
|
-Supramolecule #1000: Meiothermus taiwanensis LonA protease
Supramolecule | Name: Meiothermus taiwanensis LonA protease / type: sample / ID: 1000 / Oligomeric state: One homohexamer of MtaLonA in open form / Number unique components: 1 |
---|---|
Molecular weight | Experimental: 540 KDa / Theoretical: 540 KDa / Method: Sedimentation |
-Macromolecule #1: Meiothermus taiwanensis LonA protease
Macromolecule | Name: Meiothermus taiwanensis LonA protease / type: protein_or_peptide / ID: 1 / Name.synonym: MtaLonA / Oligomeric state: Hexamer / Recombinant expression: Yes |
---|---|
Source (natural) | Organism: Meiothermus taiwanensis (bacteria) / synonym: Meiothermus taiwanensis |
Molecular weight | Experimental: 540 KDa / Theoretical: 540 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant plasmid: pET21a |
-Experimental details
-Structure determination
Method | negative staining, cryo EM |
---|---|
Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.12 mg/mL |
---|---|
Buffer | pH: 8 / Details: 20mM Tris-HCl, 100mM NaCl, 1mM DTT, 15mM MgCl2 |
Staining | Type: NEGATIVE Details: Grids with adsorbed protein floated on 2% w/v uranyl acetate for 60 seconds. |
Grid | Details: 200 mesh gold grid with thin carbon support, glow discharged in amylamine atmosphere |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV Method: 1. wait for 10 seconds before blot 2. Blot for 3 seconds before plunging |
-Electron microscopy
Microscope | FEI TECNAI 20 |
---|---|
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification |
Date | Nov 27, 2013 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 192 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 85600 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 3.35 µm / Nominal defocus min: 1.625 µm / Nominal magnification: 62000 |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
-Image processing
Details | The particle images were interactively selected using EMAN Boxer program |
---|---|
CTF correction | Details: Each particle |
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 12.6 Å / Resolution method: OTHER / Software - Name: EMAN2 / Number images used: 30249 |
Final two d classification | Number classes: 80 |