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- PDB-5ofq: Crystal structure of substrate-free CYP109A2 from Bacillus megaterium -

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Basic information

Entry
Database: PDB / ID: 5ofq
TitleCrystal structure of substrate-free CYP109A2 from Bacillus megaterium
ComponentsCytochrome P450
KeywordsOXIDOREDUCTASE / Bacterial Proteins / Bacillus Megaterium / Bacillus / Cytochrome P-450 Enzyme System / Cytochrome P450 / Hydroxylation / Heme / Oxidation-Reduction / 25-Hydroxyvitamin D3 / Biocatalysis / Calcifediol / Cholecalciferol
Function / homology
Function and homology information


Oxidoreductases; Acting on paired donors, with incorporation or reduction of molecular oxygen / oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen / monooxygenase activity / iron ion binding / heme binding
Similarity search - Function
Cytochrome P450, B-class / 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 / Cytochrome P450
Similarity search - Component
Biological speciesBacillus megaterium (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 2.7 Å
AuthorsJozwik, I.K. / Thunnissen, A.M.W.H.
Funding support Netherlands, 1items
OrganizationGrant numberCountry
European Commission289217 Netherlands
CitationJournal: FEBS J. / Year: 2017
Title: Biochemical and structural characterization of CYP109A2, a vitamin D3 25-hydroxylase from Bacillus megaterium.
Authors: Abdulmughni, A. / Jozwik, I.K. / Brill, E. / Hannemann, F. / Thunnissen, A.W.H. / Bernhardt, R.
History
DepositionJul 11, 2017Deposition site: PDBE / Processing site: PDBE
Revision 1.0Sep 27, 2017Provider: repository / Type: Initial release
Revision 1.1Oct 4, 2017Group: Database references / Category: citation / citation_author
Item: _citation.journal_abbrev / _citation.journal_id_ISSN ..._citation.journal_abbrev / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name
Revision 1.2Nov 29, 2017Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.3Jan 24, 2018Group: Source and taxonomy / Category: entity_src_gen
Item: _entity_src_gen.pdbx_host_org_ncbi_taxonomy_id / _entity_src_gen.pdbx_host_org_scientific_name
Revision 1.4Jan 17, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_ncs_dom_lim
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id / _struct_ncs_dom_lim.beg_label_comp_id / _struct_ncs_dom_lim.beg_label_seq_id / _struct_ncs_dom_lim.end_auth_comp_id / _struct_ncs_dom_lim.end_label_asym_id / _struct_ncs_dom_lim.end_label_comp_id / _struct_ncs_dom_lim.end_label_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Cytochrome P450
B: Cytochrome P450
C: Cytochrome P450
D: Cytochrome P450
hetero molecules


Theoretical massNumber of molelcules
Total (without water)191,89116
Polymers187,9464
Non-polymers3,94612
Water27015
1
A: Cytochrome P450
hetero molecules


Theoretical massNumber of molelcules
Total (without water)48,2726
Polymers46,9861
Non-polymers1,2855
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
2
B: Cytochrome P450
hetero molecules


Theoretical massNumber of molelcules
Total (without water)47,9374
Polymers46,9861
Non-polymers9513
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
3
C: Cytochrome P450
hetero molecules


Theoretical massNumber of molelcules
Total (without water)48,0794
Polymers46,9861
Non-polymers1,0933
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
4
D: Cytochrome P450
hetero molecules


Theoretical massNumber of molelcules
Total (without water)47,6032
Polymers46,9861
Non-polymers6161
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
Unit cell
Length a, b, c (Å)77.125, 155.534, 158.151
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
11(chain A and (resseq 17:18 or resseq 20:21 or resseq...
21(chain B and (resseq 17:18 or resseq 20:21 or resseq...
31(chain C and (resseq 17:18 or resseq 20:21 or resseq...
41(chain D and (resseq 17:18 or resseq 20:21 or resseq...

NCS domain segments:

Ens-ID: 1

Dom-IDComponent-IDBeg auth comp-IDBeg label comp-IDEnd auth comp-IDEnd label comp-IDSelection detailsAuth asym-IDLabel asym-IDAuth seq-IDLabel seq-ID
11PROPROMETMET(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA17 - 1817 - 18
12GLUGLUILEILE(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA20 - 2120 - 21
13SERSERVALVAL(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA23 - 2423 - 24
14GLNGLNGLNGLN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA26 - 3926 - 39
15ALAALATYRTYR(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA41 - 5741 - 57
16THRTHRLEULEU(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA59 - 6459 - 64
17LYSLYSLYSLYS(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA6565
18ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
19ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
110ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
111ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
112ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
113ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
114ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
115ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
116ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
117ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
118ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
119ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
120ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
121ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
122ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
123ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
124ILEILEASNASN(chain A and (resseq 17:18 or resseq 20:21 or resseq...AA16 - 40116 - 401
21PROPROMETMET(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB17 - 1817 - 18
22GLUGLUILEILE(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB20 - 2120 - 21
23SERSERVALVAL(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB23 - 2423 - 24
24GLNGLNGLNGLN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB26 - 3926 - 39
25ALAALATYRTYR(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB41 - 5741 - 57
26THRTHRLEULEU(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB59 - 6459 - 64
27LYSLYSLYSLYS(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB6565
28ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
29ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
210ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
211ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
212ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
213ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
214ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
215ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
216ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
217ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
218ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
219ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
220ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
221ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
222ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
223ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
224ILEILEASNASN(chain B and (resseq 17:18 or resseq 20:21 or resseq...BB16 - 40116 - 401
31PROPROMETMET(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC17 - 1817 - 18
32GLUGLUILEILE(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC20 - 2120 - 21
33SERSERVALVAL(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC23 - 2423 - 24
34GLNGLNGLNGLN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC26 - 3926 - 39
35ALAALATYRTYR(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC41 - 5741 - 57
36THRTHRLEULEU(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC59 - 6459 - 64
37LYSLYSLYSLYS(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC6565
38ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
39ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
310ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
311ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
312ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
313ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
314ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
315ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
316ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
317ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
318ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
319ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
320ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
321ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
322ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
323ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
324ILEILEASNASN(chain C and (resseq 17:18 or resseq 20:21 or resseq...CC16 - 40116 - 401
41PROPROMETMET(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD17 - 1817 - 18
42GLUGLUILEILE(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD20 - 2120 - 21
43SERSERVALVAL(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD23 - 2423 - 24
44GLNGLNGLNGLN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD26 - 3926 - 39
45ALAALATYRTYR(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD41 - 5741 - 57
46THRTHRLEULEU(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD59 - 6459 - 64
47LYSLYSLYSLYS(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD6565
48LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
49LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
410LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
411LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
412LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
413LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
414LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
415LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
416LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
417LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
418LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
419LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
420LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
421LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
422LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
423LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401
424LEULEUASNASN(chain D and (resseq 17:18 or resseq 20:21 or resseq...DD15 - 40115 - 401

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Components

#1: Protein
Cytochrome P450 /


Mass: 46986.395 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus megaterium (strain DSM 319) (bacteria)
Gene: BMD_2035 / Plasmid: pET17b / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): C43
References: UniProt: D5DF88, Oxidoreductases; Acting on paired donors, with incorporation or reduction of molecular oxygen
#2: Chemical
ChemComp-HEM / PROTOPORPHYRIN IX CONTAINING FE / HEME / Heme B


Mass: 616.487 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C34H32FeN4O4
#3: Chemical
ChemComp-1PE / PENTAETHYLENE GLYCOL / PEG400 / Polyethylene glycol


Mass: 238.278 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C10H22O6 / Comment: precipitant*YM
#4: Chemical ChemComp-SO4 / SULFATE ION / Sulfate


Mass: 96.063 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: SO4
#5: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 15 / 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: 2.52 Å3/Da / Density % sol: 51.26 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 7.5
Details: 1.9M ammonium sulfate, 0.1M HEPES pH 7.5, 2% (v/v) PEG400

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: ESRF / Beamline: ID23-2 / Wavelength: 0.873 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Apr 7, 2014
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.873 Å / Relative weight: 1
ReflectionResolution: 2.7→52.03 Å / Num. obs: 51368 / % possible obs: 97.3 % / Redundancy: 5.6 % / Biso Wilson estimate: 48.76 Å2 / CC1/2: 0.996 / Rmerge(I) obs: 0.112 / Rpim(I) all: 0.05 / Rrim(I) all: 0.124 / Net I/σ(I): 12.4
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsCC1/2Rpim(I) allRrim(I) all% possible all
2.7-2.785.71.0250.5230.4581.12698.5
11.13-52.035.60.0260.9990.0110.02891.1

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Phasing

PhasingMethod: molecular replacement
Phasing MR
Highest resolutionLowest resolution
Rotation2.75 Å49.93 Å
Translation2.75 Å49.93 Å

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Processing

Software
NameVersionClassification
EDNAdata collection
XDSdata reduction
Aimless0.2.8data scaling
PHASER2.5.5phasing
PHENIX1.10.1_2155refinement
PDB_EXTRACT3.22data extraction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5l90

5l90


Resolution: 2.7→49.927 Å / SU ML: 0.5 / Cross valid method: FREE R-VALUE / σ(F): 1.1 / Phase error: 29.79
RfactorNum. reflection% reflection
Rfree0.2751 5009 5.1 %
Rwork0.2238 --
obs0.2264 51368 97.29 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso max: 132.67 Å2 / Biso mean: 45.9872 Å2 / Biso min: 29.6 Å2
Refinement stepCycle: final / Resolution: 2.7→49.927 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms12444 0 267 15 12726
Biso mean--48.53 38.58 -
Num. residues----1545
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.01812987
X-RAY DIFFRACTIONf_angle_d1.41817571
X-RAY DIFFRACTIONf_chiral_restr0.071886
X-RAY DIFFRACTIONf_plane_restr0.012273
X-RAY DIFFRACTIONf_dihedral_angle_d15.9434908
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDNumberRefine-IDRmsType
11A8512X-RAY DIFFRACTION7.763TORSIONAL
12B8512X-RAY DIFFRACTION7.763TORSIONAL
13C8512X-RAY DIFFRACTION7.763TORSIONAL
14D8512X-RAY DIFFRACTION7.763TORSIONAL
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 30

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
2.7-2.73070.40781700.40833190336099
2.7307-2.76280.42531810.38543130331199
2.7628-2.79650.37631710.36223133330499
2.7965-2.83190.4541980.34523079327798
2.8319-2.86910.35091430.34023215335899
2.8691-2.90840.39131720.31423110328298
2.9084-2.950.34691740.29263115328998
2.95-2.9940.351620.29173135329798
2.994-3.04080.37341840.2863126331098
3.0408-3.09060.32011790.28443129330898
3.0906-3.14390.3371570.28733088324598
3.1439-3.20110.36291410.28213168330998
3.2011-3.26260.36521500.29013157330798
3.2626-3.32920.35291640.25883123328798
3.3292-3.40160.29781670.24453135330298
3.4016-3.48070.27711950.23343060325598
3.4807-3.56770.28831700.23133100327097
3.5677-3.66420.30351700.22343127329797
3.6642-3.7720.27411770.20933098327598
3.772-3.89370.26111820.21043075325797
3.8937-4.03280.25011720.1983107327997
4.0328-4.19420.25641150.19423120323597
4.1942-4.38490.22021810.19143056323797
4.3849-4.6160.25792010.17693072327397
4.616-4.90490.1941560.17273092324896
4.9049-5.28330.19161510.17633067321896
5.2833-5.81420.21781560.19153064322096
5.8142-6.65390.24211550.18443072322796
6.6539-8.37670.20841510.16283057320895
8.3767-49.93560.19291640.16362987315194

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Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

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