+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-30988 | |||||||||
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Title | Structure of Enolase from Mycobacterium tuberculosis | |||||||||
Map data | This is the combined final map with B-factor sharpening | |||||||||
Sample |
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Keywords | METAL BINDING PROTEIN | |||||||||
Function / homology | Function and homology information phosphopyruvate hydratase / phosphopyruvate hydratase complex / phosphopyruvate hydratase activity / glycolytic process / cell surface / magnesium ion binding / extracellular region Similarity search - Function | |||||||||
Biological species | Mycobacterium tuberculosis (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.08 Å | |||||||||
Authors | Bose S / Vinothkumar KR | |||||||||
Funding support | India, 2 items
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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 | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_30988.map.gz | 117.1 MB | EMDB map data format | |
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Header (meta data) | emd-30988-v30.xml emd-30988.xml | 18.7 KB 18.7 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_30988_fsc.xml | 11.3 KB | Display | FSC data file |
Images | emd_30988.png | 79.8 KB | ||
Filedesc metadata | emd-30988.cif.gz | 6.3 KB | ||
Others | emd_30988_additional_1.map.gz emd_30988_additional_2.map.gz | 93.6 MB 93.6 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-30988 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-30988 | HTTPS FTP |
-Validation report
Summary document | emd_30988_validation.pdf.gz | 628.4 KB | Display | EMDB validaton report |
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Full document | emd_30988_full_validation.pdf.gz | 628 KB | Display | |
Data in XML | emd_30988_validation.xml.gz | 12.2 KB | Display | |
Data in CIF | emd_30988_validation.cif.gz | 16.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-30988 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-30988 | HTTPS FTP |
-Related structure data
Related structure data | 7e4xMC 6l7dC 7ckpC 7clkC 7cllC 7dlrC 7e4fC 7e51C M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_30988.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | This is the combined final map with B-factor sharpening | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.07 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Additional map: The other half-map from the final refinement
File | emd_30988_additional_1.map | ||||||||||||
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Annotation | The other half-map from the final refinement | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: One of the half-map from the final refinement
File | emd_30988_additional_2.map | ||||||||||||
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Annotation | One of the half-map from the final refinement | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Octameric Enolase enzyme
Entire | Name: Octameric Enolase enzyme |
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Components |
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-Supramolecule #1: Octameric Enolase enzyme
Supramolecule | Name: Octameric Enolase enzyme / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Mycobacterium tuberculosis (bacteria) |
Molecular weight | Theoretical: 370 KDa |
-Macromolecule #1: Enolase
Macromolecule | Name: Enolase / type: protein_or_peptide / ID: 1 / Number of copies: 8 / Enantiomer: LEVO / EC number: phosphopyruvate hydratase |
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Source (natural) | Organism: Mycobacterium tuberculosis (bacteria) |
Molecular weight | Theoretical: 45.962355 KDa |
Recombinant expression | Organism: Mycolicibacterium smegmatis (bacteria) |
Sequence | String: MHHHHHHMPI IEQVRAREIL DSRGNPTVEV EVALIDGTFA RAAVPSGAST GEHEAVELRD GGDRYGGKGV QKAVQAVLDE IGPAVIGLN ADDQRLVDQA LVDLDGTPDK SRLGGNAILG VSLAVAKAAA DSAELPLFRY VGGPNAHILP VPMMNILNGG A HADTAVDI ...String: MHHHHHHMPI IEQVRAREIL DSRGNPTVEV EVALIDGTFA RAAVPSGAST GEHEAVELRD GGDRYGGKGV QKAVQAVLDE IGPAVIGLN ADDQRLVDQA LVDLDGTPDK SRLGGNAILG VSLAVAKAAA DSAELPLFRY VGGPNAHILP VPMMNILNGG A HADTAVDI QEFMVAPIGA PSFVEALRWG AEVYHALKSV LKKEGLSTGL GDEGGFAPDV AGTTAALDLI SRAIESAGLR PG ADVALAL DAAATEFFTD GTGYVFEGTT RTADQMTEFY AGLLGAYPLV SIEDPLSEDD WDGWAALTAS IGDRVQIVGD DIF VTNPER LEEGIERGVA NALLVKVNQI GTLTETLDAV TLAHHGGYRT MISHRSGETE DTMIADLAVA IGSGQIKTGA PARS ERVAK YNQLLRIEEA LGDAARYAGD LAFPRFACET K UniProtKB: Enolase |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 3.5 mg/mL | |||||||||||||||
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Buffer | pH: 7 Component:
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Grid | Model: Quantifoil / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 90 sec. / Pretreatment - Atmosphere: AIR | |||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 289 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | TFS KRIOS |
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Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Average electron dose: 27.7 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Calibrated magnification: 130841 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal magnification: 75000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |