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- PDB-8h5m: Crystal structure of PETase S121E/D186H/N233C/S242T/N246D/S282C m... -

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Basic information

Entry
Database: PDB / ID: 8h5m
TitleCrystal structure of PETase S121E/D186H/N233C/S242T/N246D/S282C mutant from Ideonella sakaiensis
ComponentsPoly(ethylene terephthalate) hydrolase
KeywordsHYDROLASE / PET hydrolase / Ideonella sakaiensis / mutant
Function / homology
Function and homology information


acetylesterase activity / poly(ethylene terephthalate) hydrolase / carboxylic ester hydrolase activity / xenobiotic catabolic process / extracellular region
Similarity search - Function
Cutinase / PET hydrolase-like / : / Alpha/Beta hydrolase fold
Similarity search - Domain/homology
Poly(ethylene terephthalate) hydrolase
Similarity search - Component
Biological speciesIdeonella sakaiensis (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.89 Å
AuthorsLee, S.H. / Seo, H. / Kim, K.-J.
Funding support1items
OrganizationGrant numberCountry
Other government
CitationJournal: To Be Published
Title: A case of balance engineering exhibits kinetic relationship between mesophilic and thermophilic poly(ethylene terephthalate) depolymerases.
Authors: Lee, S.H. / Seo, H. / Kim, K.J.
History
DepositionOct 13, 2022Deposition site: PDBJ / Processing site: PDBE
Revision 1.0Oct 25, 2023Provider: repository / Type: Initial release
Revision 1.1Oct 9, 2024Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature / Item: _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Poly(ethylene terephthalate) hydrolase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)31,6002
Polymers31,5761
Non-polymers241
Water3,999222
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration, Size exclusion chromatography
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area90 Å2
ΔGint-7 kcal/mol
Surface area9700 Å2
MethodPISA
Unit cell
Length a, b, c (Å)50.677, 67.795, 78.471
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121

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Components

#1: Protein Poly(ethylene terephthalate) hydrolase / PET hydrolase / PETase / PET-digesting enzyme


Mass: 31576.123 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Ideonella sakaiensis (bacteria) / Gene: ISF6_4831 / Production host: Escherichia coli (E. coli)
References: UniProt: A0A0K8P6T7, poly(ethylene terephthalate) hydrolase
#2: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 222 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestN
Has protein modificationY

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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: 2.13 Å3/Da / Density % sol: 42.38 %
Crystal growTemperature: 295 K / Method: vapor diffusion, sitting drop
Details: 20 % (W/V) PEG 8K 0.1 M MES/sodium hydroxide pH 8.0, 0.2 M Calcium acetate

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: PAL/PLS / Beamline: 7A (6B, 6C1) / Wavelength: 0.97934 Å
DetectorType: ADSC QUANTUM 270 / Detector: CCD / Date: May 12, 2020
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97934 Å / Relative weight: 1
ReflectionResolution: 1.89→50 Å / Num. obs: 21162 / % possible obs: 94.6 % / Redundancy: 4.3 % / Rmerge(I) obs: 0.097 / Rpim(I) all: 0.049 / Rrim(I) all: 0.109 / Χ2: 2.416 / Net I/σ(I): 13.3 / Num. measured all: 91143
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsNum. unique obsCC1/2Rpim(I) allRrim(I) allΧ2% possible all
1.89-1.923.20.34410000.6210.2130.4071.65790.6
1.92-1.963.40.32510150.7090.1950.3821.76491.8
1.96-23.70.28610000.8190.1640.3321.88293.3
2-2.043.70.27410370.8310.1580.3191.93293.1
2.04-2.0840.25410090.8350.1420.2932.02794.5
2.08-2.133.90.22610540.90.1270.2612.01994.6
2.13-2.184.20.19910510.9230.1070.2282.06894.7
2.18-2.244.20.18810390.9320.1010.2152.08995.7
2.24-2.314.10.1510500.9620.080.1712.00493.3
2.31-2.384.10.16810310.9620.090.1922.19995.4
2.38-2.4740.12910530.9650.0690.1472.12395
2.47-2.5740.1210400.9730.0640.1372.20893
2.57-2.684.20.10910520.9810.0560.1232.30194.3
2.68-2.824.20.10110420.9860.0520.1142.41792.8
2.82-34.30.0910460.990.0450.1012.59694.1
3-3.234.90.08210870.9920.0390.0912.8196.4
3.23-3.565.30.07411030.9940.0340.0822.97597.8
3.56-4.075.40.06611090.9960.0290.0723.22597.2
4.07-5.135.30.05911440.9960.0270.0653.17898.1
5.13-505.20.05812000.9960.0270.0642.81996.7

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Processing

Software
NameVersionClassification
REFMAC5.8.0352refinement
HKL-2000data scaling
PDB_EXTRACT3.27data extraction
HKL-2000data reduction
MOLREPphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5XJH

5xjh


Resolution: 1.89→28.23 Å / Cor.coef. Fo:Fc: 0.959 / Cor.coef. Fo:Fc free: 0.94 / SU B: 2.594 / SU ML: 0.076 / Cross valid method: THROUGHOUT / σ(F): 0 / ESU R: 0.129 / ESU R Free: 0.121 / Stereochemistry target values: MAXIMUM LIKELIHOOD
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS U VALUES : REFINED INDIVIDUALLY
RfactorNum. reflection% reflectionSelection details
Rfree0.1884 1130 5.3 %RANDOM
Rwork0.151 ---
obs0.153 20017 94.65 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso max: 55.31 Å2 / Biso mean: 14.068 Å2 / Biso min: 6.26 Å2
Baniso -1Baniso -2Baniso -3
1--1.38 Å2-0 Å2-0 Å2
2--0.27 Å2-0 Å2
3---1.11 Å2
Refinement stepCycle: final / Resolution: 1.89→28.23 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1921 0 1 222 2144
Biso mean--6.26 27.1 -
Num. residues----261
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0090.0111985
X-RAY DIFFRACTIONr_bond_other_d0.0010.0161717
X-RAY DIFFRACTIONr_angle_refined_deg1.5081.6362706
X-RAY DIFFRACTIONr_angle_other_deg0.5071.5464018
X-RAY DIFFRACTIONr_dihedral_angle_1_deg6.7265264
X-RAY DIFFRACTIONr_dihedral_angle_2_deg18.0891013
X-RAY DIFFRACTIONr_dihedral_angle_3_deg13.75210292
X-RAY DIFFRACTIONr_chiral_restr0.0740.2297
X-RAY DIFFRACTIONr_gen_planes_refined0.0090.022330
X-RAY DIFFRACTIONr_gen_planes_other0.0010.02398
LS refinement shellResolution: 1.89→1.937 Å / Rfactor Rfree error: 0
RfactorNum. reflection% reflection
Rfree0.265 81 -
Rwork0.204 1360 -
obs--89.23 %

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