[English] 日本語
![](img/lk-miru.gif)
- PDB-6mw3: EM structure of Bacillus subtilis ribonucleotide reductase inhibi... -
+
Open data
-
Basic information
Entry | Database: PDB / ID: 6mw3 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | EM structure of Bacillus subtilis ribonucleotide reductase inhibited filament composed of NrdE alpha subunit and NrdF beta subunit with dATP | |||||||||
![]() |
| |||||||||
![]() | OXIDOREDUCTASE / ribonucleotide reductase / allostery / nucleotide metabolism / filament / dATP / ATP | |||||||||
Function / homology | ![]() ribonucleoside-diphosphate reductase complex / ribonucleoside-diphosphate reductase / ribonucleoside-diphosphate reductase activity, thioredoxin disulfide as acceptor / deoxyribonucleotide biosynthetic process / DNA replication / ATP binding Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | ELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 4.65 Å | |||||||||
![]() | Thomas, W.C. / Bacik, J.P. / Kaelber, J.T. / Ando, N. | |||||||||
Funding support | ![]()
| |||||||||
![]() | ![]() Title: Convergent allostery in ribonucleotide reductase. Authors: William C Thomas / F Phil Brooks / Audrey A Burnim / John-Paul Bacik / JoAnne Stubbe / Jason T Kaelber / James Z Chen / Nozomi Ando / ![]() Abstract: Ribonucleotide reductases (RNRs) use a conserved radical-based mechanism to catalyze the conversion of ribonucleotides to deoxyribonucleotides. Within the RNR family, class Ib RNRs are notable for ...Ribonucleotide reductases (RNRs) use a conserved radical-based mechanism to catalyze the conversion of ribonucleotides to deoxyribonucleotides. Within the RNR family, class Ib RNRs are notable for being largely restricted to bacteria, including many pathogens, and for lacking an evolutionarily mobile ATP-cone domain that allosterically controls overall activity. In this study, we report the emergence of a distinct and unexpected mechanism of activity regulation in the sole RNR of the model organism Bacillus subtilis. Using a hypothesis-driven structural approach that combines the strengths of small-angle X-ray scattering (SAXS), crystallography, and cryo-electron microscopy (cryo-EM), we describe the reversible interconversion of six unique structures, including a flexible active tetramer and two inhibited helical filaments. These structures reveal the conformational gymnastics necessary for RNR activity and the molecular basis for its control via an evolutionarily convergent form of allostery. | |||||||||
History |
|
-
Structure visualization
Movie |
![]() |
---|---|
Structure viewer | Molecule: ![]() ![]() |
-
Downloads & links
-
Download
PDBx/mmCIF format | ![]() | 252.6 KB | Display | ![]() |
---|---|---|---|---|
PDB format | ![]() | 204.8 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
---|
-Related structure data
Related structure data | ![]() 9272MC ![]() 9293C ![]() 6mt9C ![]() 6mv9C ![]() 6mveC ![]() 6myxC C: citing same article ( M: map data used to model this data |
---|---|
Similar structure data |
-
Links
-
Assembly
Deposited unit | ![]()
|
---|---|
1 | ![]()
| x 6
2 |
|
Symmetry | Helical symmetry: (Circular symmetry: 1 / Dyad axis: yes / N subunits divisor: 1 / Num. of operations: 6 / Rise per n subunits: 73.8 Å / Rotation per n subunits: 88.6 °) |
-
Components
#1: Protein | Mass: 80791.469 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() References: UniProt: A0A162Q3J9, UniProt: P50620*PLUS, ribonucleoside-diphosphate reductase #2: Protein/peptide | Mass: 698.854 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Details: C-terminus modeled as a polyalanine chain / Source: (gene. exp.) ![]() ![]() ![]() ![]() #3: Chemical | ChemComp-DTP / Sequence details | B. subtilis Ribonucleoside-diphosphate reductase beta subunit (NrdF) is modeled as an 8-residue ...B. subtilis Ribonucleoside-diphosphate reductase beta subunit (NrdF) is modeled as an 8-residue polyA chain. The complete sequence is provided below: MGSSHHHHHH | |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: FILAMENT / 3D reconstruction method: helical reconstruction |
-
Sample preparation
Component | Name: Inhibited filament of ribonucleoside-diphosphate reductase composed of NrdE alpha subunits and NrdF beta subunit tails Type: COMPLEX Details: Beta subunit core density only visible at low threshold. Beta subunit tail is bound with strong density to alpha subunit and modeled as a poly-A peptide in the model. Entity ID: #1-#2 / Source: RECOMBINANT | |||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Molecular weight | Experimental value: NO | |||||||||||||||||||||||||
Source (natural) | Organism: ![]() ![]() | |||||||||||||||||||||||||
Source (recombinant) | Organism: ![]() ![]() | |||||||||||||||||||||||||
Buffer solution | pH: 7.6 Details: Glycerol in original storage buffer was diluted to < 0.25% w/v. | |||||||||||||||||||||||||
Buffer component |
| |||||||||||||||||||||||||
Specimen | Conc.: 0.4 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: Cryo-EM samples of the NrdEF filament were prepared by mixing 20 uM C382S holo-NrdE with 20 or 40 uM Mn-reconstituted NrdF in assay buffer with 100 uM dATP and 1 mM CDP, prior to dilution ...Details: Cryo-EM samples of the NrdEF filament were prepared by mixing 20 uM C382S holo-NrdE with 20 or 40 uM Mn-reconstituted NrdF in assay buffer with 100 uM dATP and 1 mM CDP, prior to dilution with nucleotide-containing buffer to a concentration of 5 uM protein. A subset of the grids were pre-coated with a support film of continuous, amorphous carbon by flotation of cleaved mica. For these grids, the sample was diluted to a final protein concentration of 2 uM. | |||||||||||||||||||||||||
Specimen support | Details: unspecified | |||||||||||||||||||||||||
Vitrification | Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 95 % |
-
Electron microscopy imaging
Experimental equipment | ![]() Model: Talos Arctica / Image courtesy: FEI Company |
---|---|
Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 130000 X / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: ZEMLIN TABLEAU |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 8 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 2843 |
EM imaging optics | Energyfilter slit width: 20 eV |
Image scans | Width: 3838 / Height: 3710 |
-
Processing
EM software |
| ||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
Helical symmerty | Angular rotation/subunit: 88.6 ° / Axial rise/subunit: 73.8 Å / Axial symmetry: C1 | ||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 281591 | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.65 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 126224 / Algorithm: FOURIER SPACE / Symmetry type: HELICAL | ||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: OTHER / Space: REAL / Target criteria: Correlation coefficient | ||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 6CGN Accession code: 6CGN / Source name: PDB / Type: experimental model |