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- PDB-2wnn: Structure of wild type E. coli N-acetylneuraminic acid lyase in c... -

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Basic information

Entry
Database: PDB / ID: 2wnn
TitleStructure of wild type E. coli N-acetylneuraminic acid lyase in complex with pyruvate in space group P21
ComponentsN-ACETYLNEURAMINATE LYASE
KeywordsLYASE / CARBOHYDRATE METABOLISM
Function / homology
Function and homology information


N-acetylneuraminate lyase / N-acetylneuraminate lyase activity / N-acetylneuraminate catabolic process / single-species biofilm formation / carbohydrate metabolic process / identical protein binding / cytosol
Similarity search - Function
N-acetylneuraminate lyase / Schiff base-forming aldolase, conserved site / Dihydrodipicolinate synthase signature 1. / Schiff base-forming aldolase, active site / Dihydrodipicolinate synthase signature 2. / DapA-like / Dihydrodipicolinate synthetase family / Dihydrodipicolinate synthetase family / Aldolase class I / Aldolase-type TIM barrel ...N-acetylneuraminate lyase / Schiff base-forming aldolase, conserved site / Dihydrodipicolinate synthase signature 1. / Schiff base-forming aldolase, active site / Dihydrodipicolinate synthase signature 2. / DapA-like / Dihydrodipicolinate synthetase family / Dihydrodipicolinate synthetase family / Aldolase class I / Aldolase-type TIM barrel / TIM Barrel / Alpha-Beta Barrel / Alpha Beta
Similarity search - Domain/homology
N-acetylneuraminate lyase
Similarity search - Component
Biological speciesESCHERICHIA COLI (E. coli)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.65 Å
AuthorsCampeotto, I. / Bolt, A.H. / Harman, T.A. / Trinh, C.H. / Dennis, C.A. / Phillips, S.E.V. / Pearson, A.R. / Nelson, A. / Berry, A.
CitationJournal: J.Mol.Biol. / Year: 2010
Title: Structural Insights Into Substrate Specificity in Variants of N-Acetylneuraminic Acid Lyase Produced by Directed Evolution.
Authors: Campeotto, I. / Bolt, A.H. / Harman, T.A. / Dennis, C.A. / Trinh, C.H. / Phillips, S.E.V. / Nelson, A. / Pearson, A.R. / Berry, A.
History
DepositionJul 13, 2009Deposition site: PDBE / Processing site: PDBE
Revision 1.0Aug 25, 2010Provider: repository / Type: Initial release
Revision 1.1May 8, 2011Group: Version format compliance
Revision 1.2Jul 13, 2011Group: Version format compliance
Revision 1.3Dec 13, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Other / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_database_status / pdbx_initial_refinement_model / struct_conn / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_database_status.status_code_sf / _struct_conn.pdbx_leaving_atom_flag / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: N-ACETYLNEURAMINATE LYASE
B: N-ACETYLNEURAMINATE LYASE
C: N-ACETYLNEURAMINATE LYASE
D: N-ACETYLNEURAMINATE LYASE
hetero molecules


Theoretical massNumber of molelcules
Total (without water)135,1238
Polymers134,3854
Non-polymers7384
Water13,655758
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area10240 Å2
ΔGint-64.3 kcal/mol
Surface area39820 Å2
MethodPISA
Unit cell
Length a, b, c (Å)54.693, 142.456, 83.626
Angle α, β, γ (deg.)90.00, 109.16, 90.00
Int Tables number4
Space group name H-MP1211

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Components

#1: Protein
N-ACETYLNEURAMINATE LYASE / N-ACETYLNEURAMINIC ACID LYASE / N-ACETYLNEURAMINATE PYRUVATE-LYASE / SIALIC ACID LYASE / SIALATE ...N-ACETYLNEURAMINIC ACID LYASE / N-ACETYLNEURAMINATE PYRUVATE-LYASE / SIALIC ACID LYASE / SIALATE LYASE / SIALIC ACID ALDOLASE / NALASE


Mass: 33596.355 Da / Num. of mol.: 4 / Fragment: RESIDUES 2-296
Source method: isolated from a genetically manipulated source
Details: SCHIFF BASE BETWEEN LYS165 AND PYRUVATE IN ALL CHAINS
Source: (gene. exp.) ESCHERICHIA COLI (E. coli) / Plasmid: PKNANA / Production host: ESCHERICHIA COLI (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: P0A6L4, N-acetylneuraminate lyase
#2: Chemical ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Na
#3: Chemical ChemComp-1PE / PENTAETHYLENE GLYCOL / PEG400


Mass: 238.278 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C10H22O6 / Comment: precipitant*YM
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 758 / Source method: isolated from a natural source / Formula: H2O
Nonpolymer detailsPOLYETHYLENE GLYCOL (1PE): POLYETHYLENE GLYCOL 400 USED AS CRYOPROTECTANT, PARTIAL PEG 400 CHAINS ...POLYETHYLENE GLYCOL (1PE): POLYETHYLENE GLYCOL 400 USED AS CRYOPROTECTANT, PARTIAL PEG 400 CHAINS VISIBLE IN THE DENSITY
Sequence detailsTHE RECOMBINANT PROTEIN USED IN THIS STUDY HAS A HEXAHISTIDINE TAG RESULTING IN AN N-TERMINAL ...THE RECOMBINANT PROTEIN USED IN THIS STUDY HAS A HEXAHISTIDINE TAG RESULTING IN AN N-TERMINAL SEQUENCE OF MEHHHHHHATN INSTEAD OF THE WILD TYPE SEQUENCE MATN

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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: 2.37 Å3/Da / Density % sol: 48.07 % / Description: NONE
Crystal growpH: 8.2 / Details: 100 MM TRIS-HCL PH 8.2, 200 MM NACL, 18 % PEG 3350

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: Diamond / Beamline: I03 / Wavelength: 0.92
DetectorType: ADSC CCD / Detector: CCD / Date: May 12, 2008
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.92 Å / Relative weight: 1
Reflection twin
Crystal-IDIDOperatorDomain-IDFraction
11H,K,L10.666
11-H,-K,H+L20.334
ReflectionResolution: 1.65→79.06 Å / Num. obs: 131694 / % possible obs: 91.2 % / Observed criterion σ(I): 2 / Redundancy: 3.4 % / Biso Wilson estimate: 19.85 Å2 / Rmerge(I) obs: 0.06 / Net I/σ(I): 13.5
Reflection shellResolution: 1.65→1.74 Å / Redundancy: 2.9 % / Rmerge(I) obs: 0.32 / Mean I/σ(I) obs: 3.2 / % possible all: 60.8

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Processing

Software
NameVersionClassification
REFMAC5.5.0097refinement
MOSFLMdata reduction
SCALAdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 2WKJ

2wkj


Resolution: 1.65→79.07 Å / Cor.coef. Fo:Fc: 0.945 / Cor.coef. Fo:Fc free: 0.924 / SU B: 3.98 / SU ML: 0.121 / Cross valid method: THROUGHOUT / ESU R: 0.026 / ESU R Free: 0.026 / Stereochemistry target values: MAXIMUM LIKELIHOOD
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS. U VALUES REFINED INDIVIDUALLY. SIDE CHAINS FOR A1, A116, B3, C3, C116 WERE TRIMMED AS NO DENSITY WAS VISIBLE FOR THESE ATOMS.
RfactorNum. reflection% reflectionSelection details
Rfree0.2492 6596 5 %RANDOM
Rwork0.20986 ---
obs0.21183 125055 90.56 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.4 Å / Solvent model: MASK
Displacement parametersBiso mean: 18.189 Å2
Baniso -1Baniso -2Baniso -3
1-20.22 Å20 Å24.67 Å2
2---3.37 Å20 Å2
3----16.85 Å2
Refinement stepCycle: LAST / Resolution: 1.65→79.07 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms9120 0 31 758 9909
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0120.0229366
X-RAY DIFFRACTIONr_bond_other_d0.0010.026260
X-RAY DIFFRACTIONr_angle_refined_deg1.4041.97912681
X-RAY DIFFRACTIONr_angle_other_deg0.933.00115320
X-RAY DIFFRACTIONr_dihedral_angle_1_deg6.11251197
X-RAY DIFFRACTIONr_dihedral_angle_2_deg39.87124.902408
X-RAY DIFFRACTIONr_dihedral_angle_3_deg13.992151620
X-RAY DIFFRACTIONr_dihedral_angle_4_deg20.7051544
X-RAY DIFFRACTIONr_chiral_restr0.0790.21437
X-RAY DIFFRACTIONr_gen_planes_refined0.0070.02110460
X-RAY DIFFRACTIONr_gen_planes_other0.0010.021796
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_it0.6831.55873
X-RAY DIFFRACTIONr_mcbond_other0.1861.52431
X-RAY DIFFRACTIONr_mcangle_it1.09729422
X-RAY DIFFRACTIONr_mcangle_other
X-RAY DIFFRACTIONr_scbond_it1.73633493
X-RAY DIFFRACTIONr_scbond_other
X-RAY DIFFRACTIONr_scangle_it2.4344.53250
X-RAY DIFFRACTIONr_scangle_other
X-RAY DIFFRACTIONr_long_range_B_refined
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: 1.65→1.69 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.346 308 -
Rwork0.258 5255 -
obs--52.24 %

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