[English] 日本語
Yorodumi- EMDB-28230: E. coli 70S ribosome with A-loop mutations U2554C and U2555C: A-s... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-28230 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | E. coli 70S ribosome with A-loop mutations U2554C and U2555C: A-site tRNA class 2 | |||||||||
Map data | E. coli 70S ribosome with A-loop mutations U2554C and U2555C: A-site tRNA class 2 | |||||||||
Sample |
| |||||||||
Keywords | RNA / thermophile / A loop / RIBOSOME | |||||||||
Biological species | Escherichia coli (E. coli) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.34 Å | |||||||||
Authors | Nissley AJ / Penev PI / Watson ZL / Banfield JF / Cate JHD | |||||||||
Funding support | United States, 1 items
| |||||||||
Citation | Journal: Nucleic Acids Res / Year: 2023 Title: Rare ribosomal RNA sequences from archaea stabilize the bacterial ribosome. Authors: Amos J Nissley / Petar I Penev / Zoe L Watson / Jillian F Banfield / Jamie H D Cate / Abstract: The ribosome serves as the universally conserved translator of the genetic code into proteins and supports life across diverse temperatures ranging from below freezing to above 120°C. Ribosomes are ...The ribosome serves as the universally conserved translator of the genetic code into proteins and supports life across diverse temperatures ranging from below freezing to above 120°C. Ribosomes are capable of functioning across this wide range of temperatures even though the catalytic site for peptide bond formation, the peptidyl transferase center, is nearly universally conserved. Here we find that Thermoproteota, a phylum of thermophilic Archaea, substitute cytidine for uridine at large subunit rRNA positions 2554 and 2555 (Escherichia coli numbering) in the A loop, immediately adjacent to the binding site for the 3'-end of A-site tRNA. We show by cryo-EM that E. coli ribosomes with uridine to cytidine mutations at these positions retain the proper fold and post-transcriptional modification of the A loop. Additionally, these mutations do not affect cellular growth, protect the large ribosomal subunit from thermal denaturation, and increase the mutational robustness of nucleotides in the peptidyl transferase center. This work identifies sequence variation across archaeal ribosomes in the peptidyl transferase center that likely confers stabilization of the ribosome at high temperatures and develops a stable mutant bacterial ribosome that can act as a scaffold for future ribosome engineering efforts. | |||||||||
History |
|
-Structure visualization
Supplemental images |
---|
-Downloads & links
-EMDB archive
Map data | emd_28230.map.gz | 307.2 MB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-28230-v30.xml emd-28230.xml | 32.1 KB 32.1 KB | Display Display | EMDB header |
Images | emd_28230.png | 151.7 KB | ||
Filedesc metadata | emd-28230.cif.gz | 4.6 KB | ||
Others | emd_28230_half_map_1.map.gz emd_28230_half_map_2.map.gz | 301.9 MB 301.9 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-28230 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-28230 | HTTPS FTP |
-Validation report
Summary document | emd_28230_validation.pdf.gz | 1.2 MB | Display | EMDB validaton report |
---|---|---|---|---|
Full document | emd_28230_full_validation.pdf.gz | 1.2 MB | Display | |
Data in XML | emd_28230_validation.xml.gz | 17.3 KB | Display | |
Data in CIF | emd_28230_validation.cif.gz | 20.4 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28230 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28230 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|
-Map
File | Download / File: emd_28230.map.gz / Format: CCP4 / Size: 325 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | E. coli 70S ribosome with A-loop mutations U2554C and U2555C: A-site tRNA class 2 | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.8279 Å | ||||||||||||||||||||
Density |
| ||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
|
-Supplemental data
-Half map: E. coli 70S ribosome with A-loop mutations U2554C...
File | emd_28230_half_map_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | E. coli 70S ribosome with A-loop mutations U2554C and U2555C: A-site tRNA class 2 (half map) | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: E. coli 70S ribosome with A-loop mutations U2554C...
File | emd_28230_half_map_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | E. coli 70S ribosome with A-loop mutations U2554C and U2555C: A-site tRNA class 2 (half map) | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Sample components
-Entire : E. coli 70S ribosome with A-loop mutations U2554C and U2555C
Entire | Name: E. coli 70S ribosome with A-loop mutations U2554C and U2555C |
---|---|
Components |
|
-Supramolecule #1: E. coli 70S ribosome with A-loop mutations U2554C and U2555C
Supramolecule | Name: E. coli 70S ribosome with A-loop mutations U2554C and U2555C type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#54 |
---|---|
Source (natural) | Organism: Escherichia coli (E. coli) / Strain: BL21 |
Molecular weight | Theoretical: 850 KDa |
-Supramolecule #2: 50S Subunit
Supramolecule | Name: 50S Subunit / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#5, #27-#54 |
---|---|
Source (natural) | Organism: Escherichia coli (E. coli) / Strain: BL21 |
-Supramolecule #3: 30S Subunit
Supramolecule | Name: 30S Subunit / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #6-#26 |
---|---|
Source (natural) | Organism: Escherichia coli (E. coli) / Strain: BL21 |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.2 mg/mL | ||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Buffer | pH: 7.5 Component:
| ||||||||||||||||
Grid | Model: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 12 sec. | ||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK III |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
---|---|
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: OTHER / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 105000 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Source name: PDB / Chain - Initial model type: experimental model |
---|---|
Refinement | Space: REAL / Protocol: OTHER / Overall B value: 38.8 |