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- PDB-8ekt: CYP51 from Acanthamoeba castellanii in complex with the tetrazole... -

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Basic information

Entry
Database: PDB / ID: 8ekt
TitleCYP51 from Acanthamoeba castellanii in complex with the tetrazole-based IND inhibitor VT-1161(VT1)
Componentssterol 14a-demethylase
KeywordsOXIDOREDUCTASE/INHIBITOR / cytochrome P450 / CYP51 / sterol 14alpha-demethylase / sterol biosynthesis / OXIDOREDUCTASE-INHIBITOR complex
Function / homology
Function and homology information


oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen / methyltransferase activity / monooxygenase activity / methylation / iron ion binding / heme binding / membrane
Similarity search - Function
: / Cytochrome P450, E-class, group IV / Cytochrome p450 / Cytochrome P450 / Cytochrome P450, conserved site / Cytochrome P450 cysteine heme-iron ligand signature. / Cytochrome P450 / Cytochrome P450 superfamily / Cytochrome P450 / Orthogonal Bundle / Mainly Alpha
Similarity search - Domain/homology
PROTOPORPHYRIN IX CONTAINING FE / Chem-VT1 / Obtusifoliol 14alphademethylase, putative
Similarity search - Component
Biological speciesAcanthamoeba castellanii (eukaryote)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.29 Å
AuthorsHargrove, T.Y. / Wawrzak, Z. / Lepesheva, G.I.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM067871 United States
CitationJournal: J.Med.Chem. / Year: 2024
Title: Identification of Potent and Selective Inhibitors of Acanthamoeba : Structural Insights into Sterol 14 alpha-Demethylase as a Key Drug Target.
Authors: Hargrove, T.Y. / Lamb, D.C. / Wawrzak, Z. / Hull, M. / Kelly, S.L. / Guengerich, F.P. / Lepesheva, G.I.
History
DepositionSep 21, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 2, 2023Provider: repository / Type: Initial release
Revision 1.1May 1, 2024Group: Data collection / Database references
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.title / _citation.year
Revision 1.2May 15, 2024Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: sterol 14a-demethylase
B: sterol 14a-demethylase
C: sterol 14a-demethylase
D: sterol 14a-demethylase
E: sterol 14a-demethylase
F: sterol 14a-demethylase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)321,54718
Polymers314,6846
Non-polymers6,86312
Water8,071448
1
A: sterol 14a-demethylase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)53,5913
Polymers52,4471
Non-polymers1,1442
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
B: sterol 14a-demethylase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)53,5913
Polymers52,4471
Non-polymers1,1442
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
C: sterol 14a-demethylase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)53,5913
Polymers52,4471
Non-polymers1,1442
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
4
D: sterol 14a-demethylase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)53,5913
Polymers52,4471
Non-polymers1,1442
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
5
E: sterol 14a-demethylase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)53,5913
Polymers52,4471
Non-polymers1,1442
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
6
F: sterol 14a-demethylase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)53,5913
Polymers52,4471
Non-polymers1,1442
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)99.453, 99.304, 107.915
Angle α, β, γ (deg.)81.220, 84.080, 60.080
Int Tables number1
Space group name H-MP1

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Components

#1: Protein
sterol 14a-demethylase / Obtusifoliol 14alphademethylase / putative


Mass: 52447.324 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Acanthamoeba castellanii (eukaryote) / Plasmid: pCW / Production host: Escherichia coli (E. coli) / Strain (production host): HMS-174 / References: UniProt: L8GJB3, sterol 14alpha-demethylase
#2: Chemical
ChemComp-HEM / PROTOPORPHYRIN IX CONTAINING FE / HEME


Mass: 616.487 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C34H32FeN4O4
#3: Chemical
ChemComp-VT1 / (R)-2-(2,4-Difluorophenyl)-1,1-difluoro-3-(1H-tetrazol-1-yl)-1-(5-(4-(2,2,2-trifluoroethoxy)phenyl)pyridin-2-yl)propan-2-ol


Mass: 527.394 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C23H16F7N5O2 / Feature type: SUBJECT OF INVESTIGATION
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 448 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY

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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: 3.04 Å3/Da / Density % sol: 59 %
Crystal growTemperature: 298 K / Method: vapor diffusion, hanging drop / pH: 6.7
Details: 18% PEG8000, 0.1 M sodium citrate, 0.34 mM n-dodecyl-b-D-maltoside 5.5 mM TCEP

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 21-ID-G / Wavelength: 0.97857 Å
DetectorType: RAYONIX MX-300 / Detector: CCD / Date: Sep 16, 2022 / Details: Be lens
RadiationMonochromator: C(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97857 Å / Relative weight: 1
ReflectionResolution: 2.28→30 Å / Num. obs: 118963 / % possible obs: 98.3 % / Redundancy: 3.9 % / Biso Wilson estimate: 32.7 Å2 / CC1/2: 0.993 / Rmerge(I) obs: 0.04 / Net I/σ(I): 25
Reflection shellResolution: 2.28→2.32 Å / Rmerge(I) obs: 1.02 / Mean I/σ(I) obs: 1.7 / Num. unique obs: 1585 / CC1/2: 0.539

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Processing

Software
NameVersionClassification
REFMAC5.8.0352refinement
PDB_EXTRACT3.27data extraction
HKL-3000data reduction
HKL-3000data scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB entry 7UWP

7uwp


Resolution: 2.29→29.89 Å / Cor.coef. Fo:Fc: 0.93 / Cor.coef. Fo:Fc free: 0.921 / SU B: 6.312 / SU ML: 0.148 / Cross valid method: THROUGHOUT / σ(F): 0 / ESU R: 0.414 / ESU R Free: 0.237 / Stereochemistry target values: MAXIMUM LIKELIHOOD
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS U VALUES : REFINED INDIVIDUALLY
RfactorNum. reflection% reflectionSelection details
Rfree0.224 6472 5.2 %RANDOM
Rwork0.2126 ---
obs0.2132 118963 79 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso max: 155.93 Å2 / Biso mean: 44.581 Å2 / Biso min: 8.12 Å2
Baniso -1Baniso -2Baniso -3
1--0.01 Å20.01 Å2-0.01 Å2
2---0.01 Å2-0.01 Å2
3---0.03 Å2
Refinement stepCycle: final / Resolution: 2.29→29.89 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms19896 0 480 449 20825
Biso mean--30.07 39.29 -
Num. residues----2478
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0040.01221043
X-RAY DIFFRACTIONr_bond_other_d0.0260.01619284
X-RAY DIFFRACTIONr_angle_refined_deg0.8681.67428554
X-RAY DIFFRACTIONr_angle_other_deg0.6041.57244892
X-RAY DIFFRACTIONr_dihedral_angle_1_deg8.09252472
X-RAY DIFFRACTIONr_dihedral_angle_2_deg6.6725144
X-RAY DIFFRACTIONr_dihedral_angle_3_deg17.091103630
X-RAY DIFFRACTIONr_chiral_restr0.0410.23000
X-RAY DIFFRACTIONr_gen_planes_refined0.010.0223808
X-RAY DIFFRACTIONr_gen_planes_other0.0040.024380
LS refinement shellResolution: 2.292→2.351 Å / Rfactor Rfree error: 0 / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.319 76 -
Rwork0.294 1585 -
all-1661 -
obs--14.13 %

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