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Basic information

Entry
Database: PDB / ID: 2iuo
TitleSite Directed Mutagenesis of Key Residues Involved in the Catalytic Mechanism of Cyanase
ComponentsCYANATE HYDRATASE
KeywordsLYASE / CYANATE DEGRADATION
Function / homology
Function and homology information


cyanate catabolic process / cyanase / cyanate hydratase activity / DNA binding
Similarity search - Function
Cyanate Lyase; Chain: A, domain 2 / Cyanate lyase, C-terminal domain / : / Cyanate hydratase, N-terminal / Cyanate lyase, C-terminal / Cyanate hydratase / Cyanate lyase, C-terminal domain superfamily / Cyanate lyase C-terminal domain / Cyanate lyase C-terminal domain, Cyanate hydratase / lambda repressor-like DNA-binding domains ...Cyanate Lyase; Chain: A, domain 2 / Cyanate lyase, C-terminal domain / : / Cyanate hydratase, N-terminal / Cyanate lyase, C-terminal / Cyanate hydratase / Cyanate lyase, C-terminal domain superfamily / Cyanate lyase C-terminal domain / Cyanate lyase C-terminal domain, Cyanate hydratase / lambda repressor-like DNA-binding domains / 434 Repressor (Amino-terminal Domain) / Lambda repressor-like, DNA-binding domain superfamily / 2-Layer Sandwich / Orthogonal Bundle / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
AZIDE ION / BROMIDE ION / Cyanate hydratase
Similarity search - Component
Biological speciesESCHERICHIA COLI (E. coli)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.9 Å
AuthorsGuilloton, M. / Walsh, M.A. / Joachimiak, A. / Anderson, M.P.
Citation
Journal: To be Published
Title: A Twin Set of Low Pka Arginines Ensures the Concerted Acid Base Catalytic Mechanism of Cyanase
Authors: Guilloton, M. / Walsh, M.A. / Joachimiak, A. / Anderson, P.M.
#1: Journal: Structure / Year: 2000
Title: Structure of Cyanase Reveals that a Novel Dimeric and Decameric Arrangement of Subunits is Required for Formation of the Enzyme Active Site.
Authors: Walsh, M.A. / Otwinowski, Z. / Perrakis, A. / Anderson, P.M. / Joachimiak, A.
History
DepositionJun 6, 2006Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 8, 2006Provider: repository / Type: Initial release
Revision 1.1Feb 5, 2014Group: Database references / Derived calculations ...Database references / Derived calculations / Non-polymer description / Other / Structure summary / Version format compliance
Revision 1.2May 8, 2019Group: Data collection / Experimental preparation / Other
Category: exptl_crystal_grow / pdbx_database_proc / pdbx_database_status
Item: _exptl_crystal_grow.method / _pdbx_database_status.recvd_author_approval
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_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_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: CYANATE HYDRATASE
B: CYANATE HYDRATASE
C: CYANATE HYDRATASE
D: CYANATE HYDRATASE
E: CYANATE HYDRATASE
F: CYANATE HYDRATASE
G: CYANATE HYDRATASE
H: CYANATE HYDRATASE
I: CYANATE HYDRATASE
J: CYANATE HYDRATASE
hetero molecules


Theoretical massNumber of molelcules
Total (without water)173,78354
Polymers170,37810
Non-polymers3,40544
Water36,9672052
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area75550 Å2
ΔGint-513.1 kcal/mol
Surface area61710 Å2
MethodPQS
Unit cell
Length a, b, c (Å)76.600, 80.750, 82.280
Angle α, β, γ (deg.)70.02, 71.81, 66.34
Int Tables number1
Space group name H-MP1
Noncrystallographic symmetry (NCS)NCS oper:
IDCodeMatrixVector
1given(0.94083, -0.17133, -0.29237), (-0.2114, 0.37756, -0.90153), (0.26484, 0.91, 0.319)0.00139, -0.23242, 0.08267
2given(-0.95608, 0.17287, -0.23669), (0.16079, -0.36582, -0.91669), (-0.24505, -0.91449, 0.32196)-0.03444, -0.09693, 0.02701
3given(-0.84303, 0.50927, 0.17304), (0.51126, 0.65879, 0.55192), (0.16707, 0.55375, -0.81575)-0.07372, 0.19513, -0.14737
4given(0.84808, -0.49264, -0.19512), (-0.5094, -0.65664, -0.55618), (0.14588, 0.57108, -0.80783)0.18934, -0.10645, 0.07467
5given(-0.93274, 0.2024, 0.29839), (0.21299, -0.35846, 0.90892), (0.29092, 0.91134, 0.29124)0.04072, 0.00294, 0.10835
6given(-0.9993, 0.00547, 0.03701), (-0.00533, -0.99998, 0.00386), (0.03703, 0.00366, 0.99931)9.0E-5, 0.02518, 0.15663
7given(0.85218, -0.50683, 0.13004), (-0.48268, -0.66548, 0.56935), (-0.20202, -0.54796, -0.81174)0.1029, -0.01144, -0.13159
8given(0.94353, -0.21329, 0.25347), (-0.16556, 0.35912, 0.91849), (-0.28694, -0.90859, 0.30353)0.12908, 0.08853, -0.01976
9given(-0.85846, 0.48518, -0.16627), (0.48297, 0.65565, -0.58039), (-0.17258, -0.57855, -0.79718)-0.08016, 0.04758, -0.05508
DetailsTHE ENZYME IS A DECAMER MADE UO OF 5 DIMERS. 4 SUBUNITSCONTRIBUTE TO MAKING UP THE 5 ACTIVE SITES OF THISDECAMERIC ENZYME.THE DECAMER COULD BE VISUALIZED AS JI/CH/GE/FB/DA DIMERS.

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Components

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Protein , 1 types, 10 molecules ABCDEFGHIJ

#1: Protein
CYANATE HYDRATASE / CYANASE LYASE / CYANASE / CYANATE HYDROLASE


Mass: 17037.768 Da / Num. of mol.: 10 / Mutation: YES
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) ESCHERICHIA COLI (E. coli) / Production host: ESCHERICHIA COLI (E. coli) / Strain (production host): XL1-BLUE / References: UniProt: P00816, cyanase

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Non-polymers , 5 types, 2096 molecules

#2: Chemical
ChemComp-BR / BROMIDE ION


Mass: 79.904 Da / Num. of mol.: 10 / Source method: obtained synthetically / Formula: Br
#3: Chemical
ChemComp-CL / CHLORIDE ION


Mass: 35.453 Da / Num. of mol.: 10 / Source method: obtained synthetically / Formula: Cl
#4: Chemical...
ChemComp-SO4 / SULFATE ION


Mass: 96.063 Da / Num. of mol.: 23 / Source method: obtained synthetically / Formula: SO4
#5: Chemical ChemComp-AZI / AZIDE ION


Mass: 42.020 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: N3
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 2052 / Source method: isolated from a natural source / Formula: H2O

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Details

Sequence detailsSERINE 122 MUTATED TO GLYCINE

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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: 2.67 Å3/Da / Density % sol: 54 %
Crystal growMethod: vapor diffusion, sitting drop / pH: 7.3
Details: CRYSTALS WERE GROWN BY THE SITTING DROP METHOD OF VAPOUR DIFFUSION FROM 50% AMMONIUM SULPHATE SOLUTIONS BUFFERED WITH 50MM NAKPO4, PH = 7.3, AND IN THE PRESENCE OF 50 MM TRIC/HCL, PH =7.3.

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 19-ID / Wavelength: 1.033
DetectorType: ADSC CCD / Detector: CCD / Details: MIRRORS
RadiationMonochromator: SI 111 / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.033 Å / Relative weight: 1
ReflectionResolution: 1.85→40 Å / Num. obs: 129043 / % possible obs: 92.5 % / Redundancy: 2.2 % / Rmerge(I) obs: 0.05 / Net I/σ(I): 14
Reflection shellResolution: 1.85→1.92 Å / Redundancy: 2.1 % / Rmerge(I) obs: 0.24 / Mean I/σ(I) obs: 2.5 / % possible all: 59.9

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Processing

Software
NameVersionClassification
REFMAC5.2.0019refinement
DENZOdata reduction
SCALEPACKdata scaling
AMoREphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 1DW9

1dw9


Resolution: 1.9→76.7 Å / Cor.coef. Fo:Fc: 0.964 / Cor.coef. Fo:Fc free: 0.933 / SU B: 3.098 / SU ML: 0.093 / Cross valid method: THROUGHOUT / ESU R: 0.141 / ESU R Free: 0.142 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS.
RfactorNum. reflection% reflectionSelection details
Rfree0.208 6261 5 %RANDOM
Rwork0.149 ---
obs0.152 117862 94.8 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.4 Å / Solvent model: MASK
Displacement parametersBiso mean: 13.54 Å2
Baniso -1Baniso -2Baniso -3
1-0.77 Å20.86 Å2-1.52 Å2
2---0.68 Å2-0.1 Å2
3---0.23 Å2
Refinement stepCycle: LAST / Resolution: 1.9→76.7 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms11960 0 138 2052 14150
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0220.02212320
X-RAY DIFFRACTIONr_bond_other_d0.0020.028334
X-RAY DIFFRACTIONr_angle_refined_deg1.8452.01816699
X-RAY DIFFRACTIONr_angle_other_deg1.079320542
X-RAY DIFFRACTIONr_dihedral_angle_1_deg6.41551582
X-RAY DIFFRACTIONr_dihedral_angle_2_deg35.12224.898490
X-RAY DIFFRACTIONr_dihedral_angle_3_deg13.748152252
X-RAY DIFFRACTIONr_dihedral_angle_4_deg10.4871570
X-RAY DIFFRACTIONr_chiral_restr0.1160.21972
X-RAY DIFFRACTIONr_gen_planes_refined0.0080.0213462
X-RAY DIFFRACTIONr_gen_planes_other0.0010.022256
X-RAY DIFFRACTIONr_nbd_refined0.240.22886
X-RAY DIFFRACTIONr_nbd_other0.2130.29291
X-RAY DIFFRACTIONr_nbtor_refined0.1730.25933
X-RAY DIFFRACTIONr_nbtor_other0.0870.26539
X-RAY DIFFRACTIONr_xyhbond_nbd_refined0.1990.21707
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_metal_ion_refined
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined0.1110.28
X-RAY DIFFRACTIONr_symmetry_vdw_other0.1880.236
X-RAY DIFFRACTIONr_symmetry_hbond_refined0.2280.237
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_symmetry_metal_ion_refined
X-RAY DIFFRACTIONr_symmetry_metal_ion_other
X-RAY DIFFRACTIONr_mcbond_it1.3421.510053
X-RAY DIFFRACTIONr_mcbond_other
X-RAY DIFFRACTIONr_mcangle_it1.509212523
X-RAY DIFFRACTIONr_mcangle_other
X-RAY DIFFRACTIONr_scbond_it2.94135067
X-RAY DIFFRACTIONr_scbond_other
X-RAY DIFFRACTIONr_scangle_it3.8184.54160
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: 1.9→1.95 Å / Total num. of bins used: 20 /
RfactorNum. reflection
Rfree0.271 329
Rwork0.186 6333

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