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- PDB-7cll: Mycobacterium tubeculosis enolase in complex with 2-Phosphoglycerate -

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Basic information

Entry
Database: PDB / ID: 7cll
TitleMycobacterium tubeculosis enolase in complex with 2-Phosphoglycerate
ComponentsEnolase
KeywordsLYASE / Mycobacterium tuberculosis / enolase / complex / 2-phosphoglycerate
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
Enolase / Enolase, conserved site / Enolase, C-terminal TIM barrel domain / Enolase, N-terminal / Enolase, C-terminal TIM barrel domain / Enolase, N-terminal domain / Enolase signature. / Enolase, C-terminal TIM barrel domain / Enolase, N-terminal domain / Enolase-like, N-terminal / Enolase-like, C-terminal domain superfamily
Similarity search - Domain/homology
2-PHOSPHOGLYCERIC ACID / ACETATE ION / DI(HYDROXYETHYL)ETHER / Enolase
Similarity search - Component
Biological speciesMycobacterium tuberculosis (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.99 Å
AuthorsAhmad, M. / Jha, B. / Tiwari, S. / Pal, R.K. / Biswal, B.K.
Funding support India, 1items
OrganizationGrant numberCountry
Department of Biotechnology (DBT, India) India
CitationJournal: 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
DepositionJul 21, 2020Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jul 28, 2021Provider: repository / Type: Initial release
Revision 1.1Aug 4, 2021Group: Database references / Structure summary / Category: audit_author / citation_author
Revision 1.2Nov 8, 2023Group: Data collection / Database references
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / citation / citation_author / database_2
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _database_2.pdbx_DOI / _database_2.pdbx_database_accession
Revision 1.3Nov 29, 2023Group: Refinement description / Category: pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Enolase
B: Enolase
C: Enolase
D: Enolase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)185,66628
Polymers183,8494
Non-polymers1,81724
Water20,5551141
1
A: Enolase
B: Enolase
C: Enolase
D: Enolase
hetero molecules

A: Enolase
B: Enolase
C: Enolase
D: Enolase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)371,33356
Polymers367,6998
Non-polymers3,63448
Water1448
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation3_455-x-1,y,-z+1/21
Buried area28650 Å2
ΔGint-258 kcal/mol
Surface area97980 Å2
MethodPISA
Unit cell
Length a, b, c (Å)97.697, 197.317, 200.553
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number20
Space group name H-MC2221

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Components

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Protein , 1 types, 4 molecules ABCD

#1: Protein
Enolase / / 2-phospho-D-glycerate hydro-lyase / 2-phosphoglycerate dehydratase


Mass: 45962.355 Da / Num. of mol.: 4
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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Non-polymers , 7 types, 1165 molecules

#2: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: Mg
#3: Chemical
ChemComp-2PG / 2-PHOSPHOGLYCERIC ACID / 2-Phosphoglyceric acid


Mass: 186.057 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C3H7O7P
#4: Chemical ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL / Glycerol


Mass: 92.094 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C3H8O3
#5: Chemical
ChemComp-ACT / ACETATE ION / Acetate


Mass: 59.044 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C2H3O2
#6: Chemical ChemComp-PEG / DI(HYDROXYETHYL)ETHER / Diethylene glycol


Mass: 106.120 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C4H10O3
#7: Chemical ChemComp-CL / CHLORIDE ION / Chloride


Mass: 35.453 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Cl
#8: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 1141 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.71 Å3/Da / Density % sol: 54.67 %
Crystal growTemperature: 289 K / Method: vapor diffusion, hanging drop
Details: 20% PEG 3350, 0.2 M ammonium acetate, pH 6.5 Bis-Tris buffer

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: ESRF / Beamline: ID29 / Wavelength: 0.9775 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Nov 14, 2018
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9775 Å / Relative weight: 1
ReflectionResolution: 1.99→50 Å / Num. obs: 121176 / % possible obs: 91.8 % / Redundancy: 9.1 % / CC1/2: 0.985 / Net I/σ(I): 10.17
Reflection shellResolution: 1.99→2.02 Å / Num. unique obs: 5330 / CC1/2: 0.598

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Processing

Software
NameVersionClassification
REFMAC5.8.0222refinement
HKL-2000data reduction
HKL-2000data scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5j04

5j04


Resolution: 1.99→49.38 Å / Cor.coef. Fo:Fc: 0.957 / Cor.coef. Fo:Fc free: 0.928 / SU B: 4.725 / SU ML: 0.125 / Cross valid method: THROUGHOUT / ESU R: 0.175 / ESU R Free: 0.159 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN USED IF PRESENT IN THE INPUT
RfactorNum. reflection% reflectionSelection details
Rfree0.22051 5913 4.9 %RANDOM
Rwork0.17495 ---
obs0.17716 114574 91.19 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso mean: 28.883 Å2
Baniso -1Baniso -2Baniso -3
1--0.42 Å2-0 Å20 Å2
2--0.52 Å20 Å2
3----0.1 Å2
Refinement stepCycle: 1 / Resolution: 1.99→49.38 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms12427 0 109 1141 13677
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0120.01512738
X-RAY DIFFRACTIONr_bond_other_d
X-RAY DIFFRACTIONr_angle_refined_deg1.6031.74617318
X-RAY DIFFRACTIONr_angle_other_deg
X-RAY DIFFRACTIONr_dihedral_angle_1_deg6.73851702
X-RAY DIFFRACTIONr_dihedral_angle_2_deg30.94617.565386
X-RAY DIFFRACTIONr_dihedral_angle_3_deg12.884151582
X-RAY DIFFRACTIONr_dihedral_angle_4_deg18.4031589
X-RAY DIFFRACTIONr_chiral_restr0.1010.21709
X-RAY DIFFRACTIONr_gen_planes_refined0.0080.0219658
X-RAY DIFFRACTIONr_gen_planes_other
X-RAY DIFFRACTIONr_nbd_refined
X-RAY DIFFRACTIONr_nbd_other
X-RAY DIFFRACTIONr_nbtor_refined
X-RAY DIFFRACTIONr_nbtor_other
X-RAY DIFFRACTIONr_xyhbond_nbd_refined
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_metal_ion_refined
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined
X-RAY DIFFRACTIONr_symmetry_vdw_other
X-RAY DIFFRACTIONr_symmetry_hbond_refined
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_symmetry_metal_ion_refined
X-RAY DIFFRACTIONr_symmetry_metal_ion_other
X-RAY DIFFRACTIONr_mcbond_it2.1722.7656802
X-RAY DIFFRACTIONr_mcbond_other
X-RAY DIFFRACTIONr_mcangle_it3.114.1338492
X-RAY DIFFRACTIONr_mcangle_other
X-RAY DIFFRACTIONr_scbond_it2.9152.8985933
X-RAY DIFFRACTIONr_scbond_other
X-RAY DIFFRACTIONr_scangle_it
X-RAY DIFFRACTIONr_scangle_other
X-RAY DIFFRACTIONr_long_range_B_refined5.78640.58320344
X-RAY DIFFRACTIONr_long_range_B_other
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 1.991→2.043 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.309 436 -
Rwork0.293 7370 -
obs--80.5 %

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