PDB-6to2: Crystal structure of CYP154C5 from Nocardia farcinica in complex ...

ID or keywords:

Entry

Database: PDB / ID: 6to2

Title

Crystal structure of CYP154C5 from Nocardia farcinica in complex with 5alpha-Androstan-3-one

Components

Cytochrome P450 monooxygenase

Keywords

OXIDOREDUCTASE / Cytochrome P450 / steroid hydroxylating monooxygenase / steroid binding / oxidoreductase-substrate complex

Function / homology

Function and homology information

cholest-4-en-3-one 26-monooxygenase activity / steroid hydroxylase activity / cholesterol catabolic process / iron ion binding / heme binding
Similarity search - Function

Biological species

Nocardia farcinica IFM 10152 (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

FOURIER SYNTHESIS / Resolution: 2 Å

Authors

Rodriguez, A. / Kluenemann, T. / Blankenfeldt, W. / Schallmey, A.

Funding support

Germany, 1items

Citation

Journal: Chembiochem / Year: 2021
Title: CYP154C5 Regioselectivity in Steroid Hydroxylation Explored by Substrate Modifications and Protein Engineering*.
Authors: Bracco, P. / Wijma, H.J. / Nicolai, B. / Buitrago, J.A.R. / Klunemann, T. / Vila, A. / Schrepfer, P. / Blankenfeldt, W. / Janssen, D.B. / Schallmey, A.

History

Deposition	Dec 11, 2019	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Nov 18, 2020	Provider: repository / Type: Initial release
Revision 1.1	Mar 31, 2021	Group: Database references / Category: citation / citation_author Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.title / _citation.year / _citation_author.name
Revision 1.2	Jan 24, 2024	Group: 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 Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	6to2.cif.gz	474.6 KB	Display	PDBx/mmCIF format
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Validation report

Summary document	6to2_validation.pdf.gz	1.6 MB	Display	wwPDB validaton report
Full document	6to2_full_validation.pdf.gz	1.6 MB	Display
Data in XML	6to2_validation.xml.gz	34.9 KB	Display
Data in CIF	6to2_validation.cif.gz	51.4 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/to/6to2 ftp://data.pdbj.org/pub/pdb/validation_reports/to/6to2	HTTPS FTP

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Related structure data

Related structure data	4j6dS S: Starting model for refinement
Similar structure data	4j6c-2 4j6d-2 4gv7-2 5l1s-d 4j6d-1 4z5p-1 2yba-2 5vn4-d 4r5w-1 5l1u-d Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Similarity search - Function & homologyF&H Search Similarity search - FunctionF&H Search Similarity search - HomologyF&H Search

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Links

PDB pages	PDBj / wwPDB / NCBI
Related items in Molecule of the Month	#82 - Oct 2006 Cytochrome p450 similarity (2)

Deposited unit

A: Cytochrome P450 monooxygenase

B: Cytochrome P450 monooxygenase

hetero molecules

92.3 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: Cytochrome P450 monooxygenase

hetero molecules

defined by author
46.1 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

B: Cytochrome P450 monooxygenase

hetero molecules

defined by author
46.1 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

Unit cell

Length a, b, c (Å)	103.411, 103.411, 218.210
Angle α, β, γ (deg.)	90.000, 90.000, 120.000
Int Tables number	146
Space group name H-M	H3

#1: Protein	Cytochrome P450 monooxygenase Mass: 45219.609 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Nocardia farcinica IFM 10152 (bacteria) Gene: NFA_53110 / Plasmid: pET28a Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: Q5YNS8
#2: Chemical	ChemComp-HEM / PROTOPORPHYRIN IX CONTAINING FE / HEME Mass: 616.487 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₃₄H₃₂FeN₄O₄
#3: Chemical	ChemComp-NQ8 / (5~{S},8~{S},9~{S},10~{S},13~{S},14~{S})-10,13-dimethyl-1,2,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydrocyclopenta[a]phenanthren-3-one Mass: 274.441 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₁₉H₃₀O / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical	ChemComp-CL / CHLORIDE ION Mass: 35.453 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Cl
#5: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 503 / Source method: isolated from a natural source / Formula: H₂O
Has ligand of interest	Y

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Experiment

Experiment	Method: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

Crystal	Density Matthews: 2.48 Å³/Da / Density % sol: 50.46 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, sitting drop / pH: 6.5 Details: 0.1M BIS-TRIS pH 6.5 0.2M ammonium sulfate 25% (w/v) PEG3350

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

SLS

/ Beamline: X06SA / Wavelength: 1 Å

Detector

Type: DECTRIS EIGER X 16M / Detector: PIXEL / Date: Nov 2, 2017

Radiation

Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray

Radiation wavelength

Wavelength: 1 Å / Relative weight: 1

Reflection

Resolution: 2→82.85 Å / Num. obs: 58738 / % possible obs: 99.9 % / Redundancy: 10.5 % / CC_1/2: 0.995 / R_merge(I) obs: 0.24 / R_pim(I) all: 0.077 / R_rim(I) all: 0.252 / Net I/σ(I): 6.2 / Num. measured all: 618378 / Scaling rejects: 250

Reflection shell

Diffraction-ID: 1

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Num. measured all	Num. unique obs	CC_1/2	R_pim(I) all	R_rim(I) all	Net I/σ(I) obs	% possible all
2-2.05	9.7	2.894	41683	4306	0.547	0.967	3.055	1.2	99.9
8.94-82.85	11.3	0.108	7515	666	0.996	0.033	0.113	18.2	99.7

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Processing

Software

Name	Version	Classification
Aimless	0.7.4	data scaling
PHENIX	dev-3707	refinement
PDB_EXTRACT	3.25	data extraction
PHENIX		phasing
DIALS		data reduction

Refinement

Method to determine structure:

FOURIER SYNTHESIS
Starting model: 4j6d

4j6d

Resolution: 2→82.85 Å / SU ML: 0.27 / Cross valid method: THROUGHOUT / σ(F): 1.96 / Phase error: 28.32

	R_factor	Num. reflection	% reflection
R_free	0.2488	2989	5.09 %
R_work	0.2073	-	-
obs	0.2094	58679	99.84 %

Solvent computation

Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å

Displacement parameters

B_iso max: 128.24 Å² / B_iso mean: 41.7707 Å² / B_iso min: 18.17 Å²

Refinement step

Cycle: final / Resolution: 2→82.85 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	6270	0	248	503	7021
B_iso mean	-	-	31.43	39.76	-
Num. residues	-	-	-	-	808

LS refinement shell

Refine-ID: X-RAY DIFFRACTION / R_factor R_free error: 0 / Total num. of bins used: 21

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Num. reflection all	% reflection obs (%)
2-2.03	0.387	136	0.3132	2608	2744	99
2.03-2.07	0.318	146	0.2836	2669	2815	100
2.07-2.11	0.3321	145	0.2753	2654	2799	100
2.11-2.15	0.2854	154	0.2669	2648	2802	100
2.15-2.19	0.28	129	0.2624	2658	2787	100
2.19-2.24	0.3074	148	0.252	2638	2786	100
2.24-2.29	0.302	160	0.2435	2639	2799	100
2.29-2.35	0.2673	127	0.2339	2687	2814	100
2.35-2.41	0.2403	134	0.236	2654	2788	100
2.41-2.48	0.2994	139	0.2298	2636	2775	100
2.48-2.56	0.263	139	0.2329	2633	2772	100
2.56-2.65	0.3194	145	0.2269	2675	2820	100
2.65-2.76	0.2925	154	0.2323	2660	2814	100
2.76-2.88	0.2577	121	0.2227	2654	2775	100
2.88-3.04	0.2912	146	0.2138	2688	2834	100
3.04-3.23	0.2447	128	0.2142	2626	2754	100
3.23-3.48	0.2649	164	0.202	2644	2808	100
3.48-3.83	0.2014	143	0.1743	2656	2799	100
3.83-4.38	0.2081	150	0.1634	2647	2797	100
4.38-5.52	0.1994	145	0.1709	2653	2798	100
5.52-82.85	0.2163	136	0.1901	2663	2799	100

Refinement TLS params.

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

ID	L11 (°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)
1	2.7798	0.4298	1.8694	1.528	-0.2915	1.4882	0.0515	0.8124	-0.3126	-0.2384	-0.035	-0.3129	0.0713	0.6881	0.0103	0.3144	0.0498	0.0542	0.5384	-0.0427	0.5599	-4.0062	20.291	6.8408
2	1.1977	-0.5274	0.7033	2.3117	-0.0839	2.7306	0.025	-0.0751	0.1646	0.0807	0.0541	-0.3334	-0.2382	0.1734	-0.1083	0.2778	-0.0388	0.0577	0.2949	-0.0401	0.3946	-7.5008	34.3449	16.804
3	1.2134	0.0344	-0.1619	2.263	-0.1137	1.0684	0.0415	-0.0723	0.0587	0.1403	-0.0171	0.0798	-0.1004	-0.1519	-0.0262	0.2105	0.0548	-0.0116	0.2541	0.0085	0.234	-30.378	23.9487	18.401
4	4.1497	-1.2087	-2.9544	3.0409	1.6064	3.2098	0.1016	0.0499	0.0575	-0.0387	-0.0457	0.2476	-0.1399	-0.2343	-0.0922	0.2969	0.0836	-0.0348	0.2912	0.0624	0.2754	-34.9733	33.3249	12.9927
5	1.2338	0.696	-0.1548	1.0616	0.243	0.6376	0.0129	-0.0143	-0.2136	0.0924	0.0195	-0.2643	0.0063	0.0317	-0.0293	0.2794	0.0457	-0.0284	0.2816	-0.0178	0.2951	-17.1276	18.7348	21.9216
6	2.1956	2.5036	-0.498	4.4472	0.4192	0.8057	-0.2011	0.1559	-0.3738	-0.2347	0.2017	-0.697	0.0637	-0.038	0.0307	0.2596	0.0332	0.0201	0.2593	0.0212	0.3379	-22.823	10.2356	10.9856
7	4.6933	-1.2006	2.842	3.4116	0.6718	2.3414	0.1617	-0.8292	-0.1266	0.436	-0.1965	0.2367	0.2857	-0.5467	0.0133	0.356	-0.0788	0.0285	0.4611	0.0165	0.521	3.8719	20.4152	-7.9326
8	1.4479	0.7622	1.2409	2.5334	0.738	3.9792	0.0408	0.0384	0.1811	-0.048	-0.0073	0.5045	-0.1886	-0.4225	-0.0302	0.2482	0.0303	0.0282	0.3208	0.0511	0.4842	7.2216	34.4743	-17.9764
9	2.2435	-0.3215	-0.6746	1.3108	0.3022	3.5016	0.1101	0.0438	0.4426	-0.0776	0.1607	0.2175	-0.3284	-0.1317	-0.249	0.3386	-0.0406	0.0235	0.2518	-0.0025	0.3627	18.8906	38.6299	-12.4657
10	1.3272	-1.7267	1.2613	7.9204	-3.9429	3.3721	-0.0376	0.2636	0.0472	-0.111	-0.2278	-0.5719	-0.158	0.4279	0.2706	0.2778	-0.0865	0.0135	0.3804	-0.0257	0.304	33.724	22.1508	-26.9425
11	0.2598	0.6304	-0.0791	2.4089	-0.2099	1.3842	0.0049	-0.0053	-0.1583	-0.0846	0.0202	-0.4052	0.0861	0.2857	-0.0225	0.2433	-0.0416	-0.016	0.3156	-0.0306	0.308	33.3291	19.7167	-11.3951
12	4.9376	0.4848	-4.4454	1.4585	-1.6052	4.9662	0.0482	-0.1436	-0.0437	0.1391	-0.0357	-0.2282	-0.2979	0.427	-0.0208	0.4112	-0.1561	-0.0026	0.3244	-0.0682	0.3846	36.159	36.9968	-16.5079
13	1.6476	-0.6551	0.0852	1.7436	-0.2932	1.0279	-0.0108	-0.0276	-0.1904	-0.1147	0.1093	0.3208	-0.0139	-0.0292	-0.0933	0.2609	-0.0451	-0.021	0.2759	0.0293	0.2597	15.4102	20.3615	-21.6168
14	0.5055	-0.2204	-0.4499	3.308	0.1591	0.7466	-0.0001	0.0543	0.0352	-0.3298	-0.0307	0.321	0.115	-0.0388	0.0209	0.3044	-0.0593	-0.0541	0.3013	0.0007	0.255	20.7455	15.1112	-27.095
15	2.229	-3.1028	-0.2781	6.4461	1.382	1.0424	-0.2065	-0.1258	-0.0019	0.1419	0.339	0.0815	0.0536	0.0919	-0.1168	0.265	-0.0246	0.0226	0.2858	0.0074	0.3819	22.8362	7.5894	-10.1669

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	chain 'A' and (resid 4 through 28 )	A	4 - 28
2	X-RAY DIFFRACTION	2	chain 'A' and (resid 29 through 71 )	A	29 - 71
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 72 through 165 )	A	72 - 165
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 166 through 230 )	A	166 - 230
5	X-RAY DIFFRACTION	5	chain 'A' and (resid 231 through 377 )	A	231 - 377
6	X-RAY DIFFRACTION	6	chain 'A' and (resid 378 through 409 )	A	378 - 409
7	X-RAY DIFFRACTION	7	chain 'B' and (resid 4 through 28 )	B	4 - 28
8	X-RAY DIFFRACTION	8	chain 'B' and (resid 29 through 71 )	B	29 - 71
9	X-RAY DIFFRACTION	9	chain 'B' and (resid 72 through 96 )	B	72 - 96
10	X-RAY DIFFRACTION	10	chain 'B' and (resid 97 through 136 )	B	97 - 136
11	X-RAY DIFFRACTION	11	chain 'B' and (resid 137 through 185 )	B	137 - 185
12	X-RAY DIFFRACTION	12	chain 'B' and (resid 186 through 230 )	B	186 - 230
13	X-RAY DIFFRACTION	13	chain 'B' and (resid 231 through 335 )	B	231 - 335
14	X-RAY DIFFRACTION	14	chain 'B' and (resid 336 through 377 )	B	336 - 377
15	X-RAY DIFFRACTION	15	chain 'B' and (resid 378 through 409 )	B	378 - 409

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