+Open data
-Basic information
Entry | Database: PDB / ID: 7e4x | |||||||||
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Title | Structure of Enolase from Mycobacterium tuberculosis | |||||||||
Components | Enolase | |||||||||
Keywords | METAL BINDING PROTEIN | |||||||||
Function / homology | Function and homology information phosphopyruvate hydratase / phosphopyruvate hydratase complex / phosphopyruvate hydratase activity / glycolytic process / magnesium ion binding / cell surface / extracellular region Similarity search - Function | |||||||||
Biological species | Mycobacterium tuberculosis (bacteria) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.08 Å | |||||||||
Authors | Bose, S. / Vinothkumar, K.R. | |||||||||
Funding support | India, 2items
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Citation | Journal: IUCrJ / Year: 2023 Title: Structural snapshots of Mycobacterium tuberculosis enolase reveal dual mode of 2PG binding and its implication in enzyme catalysis. Authors: Mohammed Ahmad / Bhavya Jha / Sucharita Bose / Satish Tiwari / Abhisek Dwivedy / Deepshikha Kar / Ravikant Pal / Richard Mariadasse / Tanya Parish / Jeyaraman Jeyakanthan / Kutti R ...Authors: Mohammed Ahmad / Bhavya Jha / Sucharita Bose / Satish Tiwari / Abhisek Dwivedy / Deepshikha Kar / Ravikant Pal / Richard Mariadasse / Tanya Parish / Jeyaraman Jeyakanthan / Kutti R Vinothkumar / Bichitra Kumar Biswal / Abstract: Enolase, a ubiquitous enzyme, catalyzes the reversible conversion of 2-phosphoglycerate (2PG) to phosphoenolpyruvate (PEP) in the glycolytic pathway of organisms of all three domains of life. The ...Enolase, a ubiquitous enzyme, catalyzes the reversible conversion of 2-phosphoglycerate (2PG) to phosphoenolpyruvate (PEP) in the glycolytic pathway of organisms of all three domains of life. The underlying mechanism of the 2PG to PEP conversion has been studied in great detail in previous work, however that of the reverse reaction remains to be explored. Here we present structural snapshots of Mycobacterium tuberculosis (Mtb) enolase in apo, PEP-bound and two 2PG-bound forms as it catalyzes the conversion of PEP to 2PG. The two 2PG-bound complex structures differed in the conformation of the bound product (2PG) viz the widely reported canonical conformation and a novel binding pose, which we refer to here as the alternate conformation. Notably, we observed two major differences compared with the forward reaction: the presence of Mg is non-obligatory for the reaction and 2PG assumes an alternate conformation that is likely to facilitate its dissociation from the active site. Molecular dynamics studies and binding free energy calculations further substantiate that the alternate conformation of 2PG causes distortions in both metal ion coordination and hydrogen-bonding interactions, resulting in an increased flexibility of the active-site loops and aiding product release. Taken together, this study presents a probable mechanism involved in PEP to 2PG catalysis that is likely to be mediated by the conformational change of 2PG at the active site. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7e4x.cif.gz | 531.6 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7e4x.ent.gz | 444.4 KB | Display | PDB format |
PDBx/mmJSON format | 7e4x.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7e4x_validation.pdf.gz | 1 MB | Display | wwPDB validaton report |
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Full document | 7e4x_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 7e4x_validation.xml.gz | 92.4 KB | Display | |
Data in CIF | 7e4x_validation.cif.gz | 141.1 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/e4/7e4x ftp://data.pdbj.org/pub/pdb/validation_reports/e4/7e4x | HTTPS FTP |
-Related structure data
Related structure data | 30988MC 6l7dC 7ckpC 7clkC 7cllC 7dlrC 7e4fC 7e51C C: citing same article (ref.) M: map data used to model this data |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
#1: Protein | Mass: 45962.355 Da / Num. of mol.: 8 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Mycobacterium tuberculosis (bacteria) Gene: eno, AYJ03_005430, DSI38_18100, ERS007663_02163, ERS013471_01432, ERS024276_01771, ERS027659_01730, ERS075361_03197, ERS094182_03347, F6W99_03724, SAMEA2683035_03102 Production host: Mycolicibacterium smegmatis (bacteria) / References: UniProt: A0A0E8NV14, phosphopyruvate hydratase |
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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 |
-Sample preparation
Component | Name: Octameric Enolase enzyme / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: all / Source: RECOMBINANT | |||||||||||||||||||||||||
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Molecular weight | Value: 0.37 MDa / Experimental value: NO | |||||||||||||||||||||||||
Source (natural) | Organism: Mycobacterium tuberculosis (bacteria) | |||||||||||||||||||||||||
Source (recombinant) | Organism: Mycolicibacterium smegmatis (bacteria) / Strain: mc24517 | |||||||||||||||||||||||||
Buffer solution | pH: 7 | |||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 3.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil | |||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 289 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: TFS KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 75000 X / Calibrated magnification: 130841 X / 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 | Electron dose: 27.7 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) |
Image scans | Sampling size: 14 µm / Width: 4096 / Height: 4096 |
-Processing
Software | Name: PHENIX / Version: 1.15.2_3472: / Classification: refinement | ||||||||||||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 153748 | ||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: D4 (2x4 fold dihedral) | ||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.08 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 75126 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: OTHER / Space: REAL | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 7CLL Accession code: 7CLL / Source name: PDB / Type: experimental model | ||||||||||||||||||||||||||||||||||||||||
Refinement | Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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