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- PDB-1k2x: Crystal structure of putative asparaginase encoded by Escherichia... -

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Basic information

Entry
Database: PDB / ID: 1k2x
TitleCrystal structure of putative asparaginase encoded by Escherichia coli ybiK gene
Components(Putative L-asparaginaseAsparaginase) x 2
KeywordsHYDROLASE / Ntn Hydrolase / Asparginase / Autoproteolysis
Function / homology
Function and homology information


beta-aspartyl-peptidase / asparaginase activity / beta-aspartyl-peptidase activity / protein autoprocessing / hydrolase activity / cytoplasm
Similarity search - Function
(Glycosyl)asparaginase / Peptidase T2, asparaginase 2 / Asparaginase / Glutamine Phosphoribosylpyrophosphate, subunit 1, domain 1 / Nucleophile aminohydrolases, N-terminal / 4-Layer Sandwich / Alpha Beta
Similarity search - Domain/homology
Isoaspartyl peptidase
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.65 Å
AuthorsBorek, D. / Jaskolski, M.
Citation
Journal: Acta Crystallogr.,Sect.D / Year: 2008
Title: Crystal packing of plant-type L-asparaginase from Escherichia coli.
Authors: Michalska, K. / Borek, D. / Hernandez-Santoyo, A. / Jaskolski, M.
#1: Journal: Acta Crystallogr.,Sect.D / Year: 2000
Title: Crystallization and Preliminary Crystallographic Studies of a New L-asparaginase Encoded by the Escherichia coli Genome
Authors: Borek, D. / Jaskolski, M.
#2: Journal: Nature / Year: 1995
Title: A Protein Catalytic Framework with an N-terminal Nucleophile is Capable of Self-activation
Authors: Brannigan, J.A. / Dodson, G. / Duggleby, H.J. / Moody, P.C. / Smith, J.L. / Tomchick, D.R. / Murzin, A.G.
#3: Journal: Protein Sci. / Year: 1998
Title: Crystal Structure of Glycosylasparaginase from Flavobacterium Meningosepticum
Authors: Xuan, J. / Tarentino, A.L. / Grimwood, B.G. / Plummer Jr., T.H. / Cui, T. / Guan, C. / Van Roey, P.
#4: Journal: Nat.Struct.Mol.Biol. / Year: 1995
Title: Three-dimensional Structure of Human Lysosomal Aspartylglucosaminidase
Authors: Oinonen, C. / Tikkanen, R. / Rouvinen, J. / Peltonen, L.
#5: Journal: J.Biol.Chem. / Year: 1998
Title: Crystal Structures of Flavobacterium Glycosylasparaginase. An N-terminal Nucleophile Hydrolase Activated by Intramolecular Proteolysis
Authors: Guo, H.C. / Xu, Q. / Buckley, D. / Guan, C.
#6: Journal: Cell(Cambridge,Mass.) / Year: 1999
Title: Structural Insights into the Mechanism of Intramolecular Proteolysis
Authors: Xu, Q. / Buckley, D. / Guan, C. / Guo, H.C.
History
DepositionSep 30, 2001Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 9, 2003Provider: repository / Type: Initial release
Revision 1.1Mar 24, 2008Group: Version format compliance
Revision 1.2Jul 13, 2011Group: Advisory / Derived calculations / Version format compliance
Revision 1.3Aug 16, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / diffrn_source / pdbx_initial_refinement_model / 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 / _diffrn_source.pdbx_synchrotron_site / _struct_conn.pdbx_leaving_atom_flag / _struct_ref_seq_dif.details / _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: Putative L-asparaginase
B: Putative L-asparaginase
C: Putative L-asparaginase
D: Putative L-asparaginase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)67,13915
Polymers66,7744
Non-polymers36511
Water10,359575
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area15440 Å2
ΔGint-193 kcal/mol
Surface area20110 Å2
MethodPISA, PQS
Unit cell
Length a, b, c (Å)50.296, 77.624, 148.152
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number19
Space group name H-MP212121
DetailsThe biological assembly is a heterotetramer generated by non-crystallographical symmetry from two alpha+beta heterodimers.

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Components

#1: Protein Putative L-asparaginase / Asparaginase / E.C.3.5.1.1 / L-asparagine Amidohydrolase


Mass: 18958.693 Da / Num. of mol.: 2 / Fragment: N-TERMINUS (RESIDUES 2-178)
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: ybiK / Plasmid: pET11d / Production host: Escherichia coli (E. coli) / Strain (production host): JM109 / References: UniProt: P37595, asparaginase
#2: Protein Putative L-asparaginase / Asparaginase / E.C.3.5.1.1 / L-asparagine Amidohydrolase


Mass: 14428.146 Da / Num. of mol.: 2 / Fragment: C-TERMINUS (RESIDUES 179-321)
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: ybiK / Plasmid: pET11d / Production host: Escherichia coli (E. coli) / Strain (production host): JM109 / References: UniProt: P37595, asparaginase
#3: Chemical ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Na
#4: Chemical
ChemComp-CL / CHLORIDE ION / Chloride


Mass: 35.453 Da / Num. of mol.: 9 / Source method: obtained synthetically / Formula: Cl
#5: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 575 / 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: 2.16 Å3/Da / Density % sol: 43.18 %
Crystal growTemperature: 292 K / Method: vapor diffusion, hanging drop / pH: 8.5
Details: PEG 4000, PEG 400, magnesium chloride, Tris-HCl pH 8.5, VAPOR DIFFUSION, HANGING DROP, temperature 292K
Crystal grow
*PLUS
Method: vapor diffusion
Components of the solutions
*PLUS
IDConc.Common nameCrystal-IDSol-IDChemical formulaDetails
10.2 M1reservoirMgCl2
20.1 MTris-HCl1reservoirpH8.5
315 %PEG4001reservoir
420 %PEG40001reservoir

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: EMBL/DESY, HAMBURG / Beamline: X31 / Wavelength: 1.0442 Å
DetectorType: MARRESEARCH / Detector: IMAGE PLATE / Date: Nov 4, 1999 / Details: Toroidal mirror
RadiationMonochromator: Double-crystal monochromator / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.0442 Å / Relative weight: 1
ReflectionResolution: 1.65→20 Å / Num. all: 70249 / Num. obs: 70249 / Observed criterion σ(F): 0 / Observed criterion σ(I): -3 / Redundancy: 5.6 % / Rmerge(I) obs: 0.075 / Net I/σ(I): 16.8
Reflection shellResolution: 1.65→1.68 Å / Redundancy: 5.5 % / Rmerge(I) obs: 0.476 / Mean I/σ(I) obs: 2.5 / % possible all: 89.9

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Processing

Software
NameVersionClassification
DENZOdata reduction
SCALEPACKdata scaling
EPMRphasing
REFMAC5refinement
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 2GAW, polyalanine model

2gaw


Resolution: 1.65→19.5 Å / SU B: 2.286 / SU ML: 0.077 / TLS residual ADP flag: LIKELY RESIDUAL / Isotropic thermal model: Isotropic / Cross valid method: THROUGHOUT / ESU R: 0.085 / ESU R Free: 0.088 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflectionSelection details
Rfree0.19836 1064 1.5 %RANDOM
Rwork0.16295 ---
obs0.16349 69077 99.52 %-
all-69077 --
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.4 Å / Solvent model: BABINET MODEL WITH MASK
Displacement parametersBiso mean: 10.18 Å2
Baniso -1Baniso -2Baniso -3
1--0.09 Å20 Å20 Å2
2--0.51 Å20 Å2
3----0.42 Å2
Refine analyze
FreeObs
Luzzati coordinate error0.088 Å0.085 Å
Refinement stepCycle: LAST / Resolution: 1.65→19.5 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms4240 0 11 575 4826
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0090.0214342
X-RAY DIFFRACTIONr_bond_other_d0.0010.024072
X-RAY DIFFRACTIONr_angle_refined_deg1.4111.9715859
X-RAY DIFFRACTIONr_angle_other_deg1.10539424
X-RAY DIFFRACTIONr_dihedral_angle_1_deg3.9923574
X-RAY DIFFRACTIONr_dihedral_angle_2_deg
X-RAY DIFFRACTIONr_dihedral_angle_3_deg15.83215760
X-RAY DIFFRACTIONr_dihedral_angle_4_deg
X-RAY DIFFRACTIONr_chiral_restr0.0730.2675
X-RAY DIFFRACTIONr_gen_planes_refined0.0040.024912
X-RAY DIFFRACTIONr_gen_planes_other0.0030.02850
X-RAY DIFFRACTIONr_nbd_refined0.30.3944
X-RAY DIFFRACTIONr_nbd_other0.2120.34204
X-RAY DIFFRACTIONr_nbtor_refined
X-RAY DIFFRACTIONr_nbtor_other0.8210.58
X-RAY DIFFRACTIONr_xyhbond_nbd_refined0.1490.5489
X-RAY DIFFRACTIONr_xyhbond_nbd_other0.1620.55
X-RAY DIFFRACTIONr_metal_ion_refined0.0990.55
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined0.1810.317
X-RAY DIFFRACTIONr_symmetry_vdw_other0.2230.345
X-RAY DIFFRACTIONr_symmetry_hbond_refined0.1870.575
X-RAY DIFFRACTIONr_symmetry_hbond_other0.080.51
X-RAY DIFFRACTIONr_mcbond_it0.5291.52850
X-RAY DIFFRACTIONr_mcangle_it1.01124508
X-RAY DIFFRACTIONr_scbond_it1.76331492
X-RAY DIFFRACTIONr_scangle_it2.9614.51352
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 1.65→1.694 Å / Total num. of bins used: 20 /
RfactorNum. reflection
Rfree0.261 79
Rwork0.252 4802
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
11.07260.2308-0.43730.4093-0.00011.65440.0182-0.20980.02450.1009-0.064-0.03050.03110.09870.04580.0497-0.01540.00220.0832-0.00630.065648.7782.54134.764
20.8390.26580.25110.6654-0.0541.41270.0331-0.18020.03050.0865-0.02780.06050.0996-0.2422-0.00530.0541-0.01980.02270.1155-0.00530.059734.0378.131132.632
30.72690.0270.08230.91240.29590.7202-0.01310.064-0.0624-0.06270.0425-0.0374-0.03740.0736-0.02940.0253-0.00340.01120.0447-0.00670.057446.09370.446102.601
40.6971-0.1248-0.07080.7340.10530.7165-0.00960.0356-0.0211-0.0507-0.01630.1038-0.0474-0.07370.02590.0241-0.0013-0.00510.0374-0.00490.056832.28476.88106.476
Refinement TLS group
IDRefine-IDRefine TLS-IDAuth asym-IDLabel asym-IDAuth seq-IDLabel seq-ID
1X-RAY DIFFRACTION1AA2 - 1571 - 156
2X-RAY DIFFRACTION2BB179 - 3131 - 135
3X-RAY DIFFRACTION3CC2 - 1571 - 156
4X-RAY DIFFRACTION4DD179 - 3131 - 135
Software
*PLUS
Version: 5 / Classification: refinement
Refinement
*PLUS
Solvent computation
*PLUS
Displacement parameters
*PLUS

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