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- PDB-7mdi: Structure of the Neisseria gonorrhoeae ribonucleotide reductase i... -
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Open data
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Basic information
Entry | Database: PDB / ID: 7mdi | |||||||||||||||
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Title | Structure of the Neisseria gonorrhoeae ribonucleotide reductase in the inactive state | |||||||||||||||
![]() | (Ribonucleoside-diphosphate reductase subunit ...) x 2 | |||||||||||||||
![]() | OXIDOREDUCTASE / inactive complex / ribonucleotide reductase | |||||||||||||||
Function / homology | ![]() 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 / single particle reconstruction / cryo EM / Resolution: 4.3 Å | |||||||||||||||
![]() | Levitz, T.S. / Drennan, C.L. / Brignole, E.J. | |||||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Effects of chameleon dispense-to-plunge speed on particle concentration, complex formation, and final resolution: A case study using the Neisseria gonorrhoeae ribonucleotide reductase inactive complex. Authors: Talya S Levitz / Edward J Brignole / Ivan Fong / Michele C Darrow / Catherine L Drennan / ![]() ![]() Abstract: Ribonucleotide reductase (RNR) is an essential enzyme that converts ribonucleotides to deoxyribonucleotides and is a promising antibiotic target, but few RNRs have been structurally characterized. We ...Ribonucleotide reductase (RNR) is an essential enzyme that converts ribonucleotides to deoxyribonucleotides and is a promising antibiotic target, but few RNRs have been structurally characterized. We present the use of the chameleon, a commercially-available piezoelectric cryogenic electron microscopy plunger, to address complex denaturation in the Neisseria gonorrhoeae class Ia RNR. Here, we characterize the extent of denaturation of the ring-shaped complex following grid preparation using a traditional plunger and using a chameleon with varying dispense-to-plunge times. We also characterize how dispense-to-plunge time influences the amount of protein sample required for grid preparation and preferred orientation of the sample. We demonstrate that the fastest dispense-to-plunge time of 54 ms is sufficient for generation of a data set that produces a high quality structure, and that a traditional plunging technique or slow chameleon dispense-to-plunge times generate data sets limited in resolution by complex denaturation. The 4.3 Å resolution structure of Neisseria gonorrhoeae class Ia RNR in the inactive α4β4 oligomeric state solved using the chameleon with a fast dispense-to-plunge time yields molecular information regarding similarities and differences to the well studied Escherichia coli class Ia RNR α4β4 ring. | |||||||||||||||
History |
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Structure visualization
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Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 722.7 KB | Display | ![]() |
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PDB format | ![]() | 564.2 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 23773MC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
-Ribonucleoside-diphosphate reductase subunit ... , 2 types, 8 molecules ABDCEGFH
#1: Protein | Mass: 87081.344 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() References: UniProt: Q5F8Z6, ribonucleoside-diphosphate reductase #2: Protein | Mass: 45107.785 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() References: UniProt: Q5F8Z5, ribonucleoside-diphosphate reductase |
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-Non-polymers , 5 types, 48 molecules ![](data/chem/img/CDP.gif)
![](data/chem/img/DTP.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/FEO.gif)
![](data/chem/img/HOH.gif)
![](data/chem/img/DTP.gif)
![](data/chem/img/MG.gif)
![](data/chem/img/FEO.gif)
![](data/chem/img/HOH.gif)
#3: Chemical | ChemComp-CDP / #4: Chemical | ChemComp-DTP / #5: Chemical | ChemComp-MG / #6: Chemical | ChemComp-FEO / #7: Water | ChemComp-HOH / | |
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-Details
Has ligand of interest | N |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Ribonucleotide reductase inactive complex / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Value: 0.528 MDa / Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: ![]() | ||||||||||||||||||||
Source (recombinant) | Organism: ![]() ![]() | ||||||||||||||||||||
Buffer solution | pH: 7.6 | ||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||
Vitrification | Cryogen name: ETHANE Details: Plunged using the chameleon (SPT labtech) Glow discharged at 12 mA for 250 s |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Talos Arctica / Image courtesy: FEI Company |
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Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 92000 X / Nominal defocus max: 3100 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 7 sec. / Electron dose: 53.15 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 620 |
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Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 55161 | ||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: D2 (2x2 fold dihedral) | ||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 50425 / Symmetry type: POINT |