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- PDB-1ovd: THE K136E MUTANT OF LACTOCOCCUS LACTIS DIHYDROOROTATE DEHYDROGENA... -

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Basic information

Entry
Database: PDB / ID: 1ovd
TitleTHE K136E MUTANT OF LACTOCOCCUS LACTIS DIHYDROOROTATE DEHYDROGENASE A IN COMPLEX WITH OROTATE
ComponentsDIHYDROOROTATE DEHYDROGENASE A
KeywordsOXIDOREDUCTASE / HOMODIMER / ALPHA-BETA BARREL / FLAVOPROTEIN / OROTATE COMPLEX / MUTANT ENZYME
Function / homology
Function and homology information


dihydroorotate dehydrogenase (fumarate) / dihydroorotate dehydrogenase (fumarate) activity / 'de novo' UMP biosynthetic process / 'de novo' pyrimidine nucleobase biosynthetic process / cytoplasm
Similarity search - Function
Dihydroorotate dehydrogenase A, chain A, domain 2 / Dihydroorotate dehydrogenase, class 1 / Dihydroorotate Dehydrogenase A; chain A, domain 2 / Dihydroorotate Dehydrogenase A, chain A, domain 2 / Dihydroorotate dehydrogenase, class 1A / Dihydroorotate dehydrogenase, class 1/ 2 / Dihydroorotate dehydrogenase signature 1. / Dihydroorotate dehydrogenase signature 2. / Dihydroorotate dehydrogenase, conserved site / Dihydroorotate dehydrogenase domain ...Dihydroorotate dehydrogenase A, chain A, domain 2 / Dihydroorotate dehydrogenase, class 1 / Dihydroorotate Dehydrogenase A; chain A, domain 2 / Dihydroorotate Dehydrogenase A, chain A, domain 2 / Dihydroorotate dehydrogenase, class 1A / Dihydroorotate dehydrogenase, class 1/ 2 / Dihydroorotate dehydrogenase signature 1. / Dihydroorotate dehydrogenase signature 2. / Dihydroorotate dehydrogenase, conserved site / Dihydroorotate dehydrogenase domain / Dihydroorotate dehydrogenase / Aldolase class I / Aldolase-type TIM barrel / Roll / TIM Barrel / Alpha-Beta Barrel / Mainly Beta / Alpha Beta
Similarity search - Domain/homology
FLAVIN MONONUCLEOTIDE / OROTIC ACID / Dihydroorotate dehydrogenase A (fumarate) / Dihydroorotate dehydrogenase A (fumarate)
Similarity search - Component
Biological speciesLactococcus lactis (lactic acid bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.25 Å
AuthorsNorager, S. / Arent, S. / Bjornberg, O. / Ottosen, M. / Lo Leggio, L. / Jensen, K.F. / Larsen, S.
Citation
Journal: J.Biol.Chem. / Year: 2003
Title: Lactococcus lactis dihydroorotate dehydrogenase A mutants reveal important facets of the enzymatic function.
Authors: Norager, S. / Arent, S. / Bjornberg, O. / Ottesen, M. / Lo Leggio, L. / Jensen, K.F. / Larsen, S.
#1: Journal: Biochemistry / Year: 1997
Title: Active Site of Dihydroorotate Dehydrogenase A from Lactococcus Lactis Investigated by Chemical Modification and Mutagenesis
Authors: Bjornberg, O. / Rowland, P. / Larsen, S. / Jensen, K.F.
#2: Journal: Protein Sci. / Year: 1998
Title: The Crystals Structure of Lactococcus Lactis Dihydroorotate Dehydrogenase A Complexed with the Enzyme Reaction Product Throws Light on its Enzymatic Function
Authors: Rowland, P. / Bjornberg, O. / Nielsen, F.S. / Jensen, K.F. / Larsen, S.
#3: Journal: Structure / Year: 1997
Title: The Crystals Structure of the Flavin Containing Enzyme Dihydroorotate Dehydrogenase A from Lactococcus Lactis
Authors: Rowland, P. / Nielsen, F.S. / Jensen, K.F. / Larsen, S.
#4: Journal: Protein Sci. / Year: 1996
Title: Purification and Characterisation of Dihydroorotate Dehydrogenase A from Lactococcus Lactis, Crystallisation and Preliminary X-Ray Diffraction Studies of the Enzyme
Authors: Nielsen, F.S. / Rowland, P. / Larsen, S. / Jensen, K.F.
History
DepositionMar 26, 2003Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 19, 2003Provider: repository / Type: Initial release
Revision 1.1Apr 29, 2008Group: Version format compliance
Revision 1.2Jul 13, 2011Group: Non-polymer description / Version format compliance
Revision 1.3Mar 7, 2018Group: Data collection / Category: diffrn_source / Item: _diffrn_source.pdbx_synchrotron_site
Revision 1.4Oct 27, 2021Group: Database references / Derived calculations
Category: database_2 / pdbx_struct_conn_angle ...database_2 / pdbx_struct_conn_angle / struct_conn / struct_ref_seq_dif / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _pdbx_struct_conn_angle.value / _struct_conn.pdbx_dist_value / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_conn.ptnr2_label_seq_id / _struct_ref_seq_dif.details / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id
Revision 1.5Aug 16, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: DIHYDROOROTATE DEHYDROGENASE A
B: DIHYDROOROTATE DEHYDROGENASE A
hetero molecules


Theoretical massNumber of molelcules
Total (without water)70,21813
Polymers68,4842
Non-polymers1,73411
Water8,773487
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area8530 Å2
ΔGint-48 kcal/mol
Surface area21710 Å2
MethodPISA
Unit cell
Length a, b, c (Å)52.870, 108.290, 65.940
Angle α, β, γ (deg.)90.00, 103.85, 90.00
Int Tables number4
Space group name H-MP1211
DetailsThe asymmetric unit contains the biological homodimer

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Components

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Protein , 1 types, 2 molecules AB

#1: Protein DIHYDROOROTATE DEHYDROGENASE A / Dihydroorotate oxidase A / DHOdehase A / DHODase A / DHOD A


Mass: 34242.102 Da / Num. of mol.: 2 / Mutation: K136E
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lactococcus lactis (lactic acid bacteria)
Gene: PYRDA / Plasmid: PUHE23 / Production host: Escherichia coli (E. coli) / Strain (production host): SO6645
References: UniProt: P54321, UniProt: A2RJT9*PLUS, EC: 1.3.3.1

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Non-polymers , 5 types, 498 molecules

#2: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#3: Chemical ChemComp-FMN / FLAVIN MONONUCLEOTIDE / RIBOFLAVIN MONOPHOSPHATE / Flavin mononucleotide


Mass: 456.344 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C17H21N4O9P
#4: Chemical ChemComp-ORO / OROTIC ACID / Orotic acid


Mass: 156.096 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C5H4N2O4
#5: Chemical
ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL / Glycerol


Mass: 92.094 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C3H8O3
#6: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 487 / Source method: isolated from a natural source / Formula: H2O

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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: 2.41 Å3/Da / Density % sol: 48.61 %
Crystal growTemperature: 293 K / Method: vapor diffusion, hanging drop / pH: 8.5
Details: PEG 6K, Sodium acetate, Tris-HCl, pH 8.5, VAPOR DIFFUSION, HANGING DROP, temperature 293K
Crystal grow
*PLUS
pH: 7 / Method: vapor diffusion
Components of the solutions
*PLUS
IDConc.Common nameCrystal-IDSol-IDDetailsChemical formula
11 mg/mlprotein1drop
20.05 Msodium phosphate1droppH7.0
325 mM1reservoirpH6.0Na2HPO4
410 %glycerol1reservoir

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: MAX II / Beamline: I711 / Wavelength: 1.098 Å
DetectorType: MARRESEARCH / Detector: CCD / Date: Sep 26, 2001 / Details: Bent mirror
RadiationMonochromator: Si(III) single crystal / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.098 Å / Relative weight: 1
ReflectionResolution: 2.25→12 Å / Num. all: 34158 / Num. obs: 33990 / % possible obs: 100 % / Observed criterion σ(F): 0 / Observed criterion σ(I): -3 / Redundancy: 4.3 % / Biso Wilson estimate: 6.2 Å2 / Rmerge(I) obs: 0.089 / Net I/σ(I): 15.3
Reflection shellResolution: 2.25→2.29 Å / Rmerge(I) obs: 0.282 / Mean I/σ(I) obs: 4.93 / Num. unique all: 1709 / % possible all: 100
Reflection
*PLUS
Num. obs: 34158 / % possible obs: 100 % / Num. measured all: 146256
Reflection shell
*PLUS
% possible obs: 100 % / Mean I/σ(I) obs: 4.9

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Processing

Software
NameClassification
CNSrefinement
DENZOdata reduction
SCALEPACKdata scaling
AMoREphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: Lactococcus lactis DHODA PDB ID 1DOR

1dor


Resolution: 2.25→12 Å / Rfactor Rfree error: 0.005 / Data cutoff high absF: 156760.05 / Data cutoff high rms absF: 156760.05 / Data cutoff low absF: 0 / Isotropic thermal model: isotropic / Cross valid method: THROUGHOUT / σ(F): 0 / Stereochemistry target values: ENGH & HUBER
RfactorNum. reflection% reflectionSelection details
Rfree0.202 1553 10 %Random
Rwork0.178 ---
obs-33990 99.9 %-
Solvent computationSolvent model: FLAT / Bsol: 42.7672 Å2 / ksol: 0.353933 e/Å3
Displacement parametersBiso mean: 19.5 Å2
Baniso -1Baniso -2Baniso -3
1-2.45 Å22.98 Å20 Å2
2--2.45 Å20 Å2
3----4.89 Å2
Refine analyze
FreeObs
Luzzati coordinate error0.25 Å0.22 Å
Luzzati d res low-5 Å
Luzzati sigma a0.2 Å0.16 Å
Refinement stepCycle: LAST / Resolution: 2.25→12 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms4818 0 116 487 5421
Refine LS restraints
Refine-IDTypeDev idealDev ideal target
X-RAY DIFFRACTIONc_bond_d0.007
X-RAY DIFFRACTIONc_angle_d1.3
X-RAY DIFFRACTIONc_dihedral_angle_d22.1
X-RAY DIFFRACTIONc_improper_angle_d0.85
X-RAY DIFFRACTIONc_mcbond_it2.271.5
X-RAY DIFFRACTIONc_scbond_it3.712
X-RAY DIFFRACTIONc_mcangle_it3.342
X-RAY DIFFRACTIONc_scangle_it5.062.5
LS refinement shellResolution: 2.25→2.39 Å / Rfactor Rfree error: 0.015 / Total num. of bins used: 10
RfactorNum. reflection% reflection
Rfree0.241 264 10 %
Rwork0.204 1383 -
obs-5642 99.9 %
Xplor file
Refine-IDSerial noParam fileTopol file
X-RAY DIFFRACTION1protein_rep.paramprotein.top
X-RAY DIFFRACTION2water_rep.paramwater.top
X-RAY DIFFRACTION3ion.param
Refine LS restraints
*PLUS
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONc_angle_d
X-RAY DIFFRACTIONc_angle_deg1.5
X-RAY DIFFRACTIONc_dihedral_angle_d
X-RAY DIFFRACTIONc_dihedral_angle_deg22.1
X-RAY DIFFRACTIONc_improper_angle_d
X-RAY DIFFRACTIONc_improper_angle_deg0.85

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