PDB-5dyz: Crystal structure of Asp251Gly/Gln307His mutant of cytochrome P45...

ID or keywords:

Entry

Database: PDB / ID: 5dyz

Title

Crystal structure of Asp251Gly/Gln307His mutant of cytochrome P450 BM3 in complex with N-palmitoylglycine

Components

Bifunctional P-450/NADPH-P450 reductase

Keywords

OXIDOREDUCTASE / cytochrome P450 / random mutagenesis / drug metabolism / substrate

Function / homology

Function and homology information

NADPH-hemoprotein reductase / NADPH-hemoprotein reductase activity / oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen, reduced flavin or flavoprotein as one donor, and incorporation of one atom of oxygen / unspecific monooxygenase / aromatase activity / metabolic process / response to hormone / FMN binding / flavin adenine dinucleotide binding / iron ion binding ...
Similarity search - Function

Biological species

Bacillus megaterium (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.967 Å

Authors

Di Nardo, G. / Dell'Angelo, V. / Gilardi, G.

Citation

Journal: Arch.Biochem.Biophys. / Year: 2016
Title: Subtle structural changes in the Asp251Gly/Gln307His P450 BM3 mutant responsible for new activity toward diclofenac, tolbutamide and ibuprofen.
Authors: Di Nardo, G. / Dell'Angelo, V. / Catucci, G. / Sadeghi, S.J. / Gilardi, G.

History

Deposition	Sep 25, 2015	Deposition site: RCSB / Processing site: PDBE
Revision 1.0	Jan 20, 2016	Provider: repository / Type: Initial release
Revision 1.1	Jun 22, 2016	Group: Database references
Revision 1.2	Jan 10, 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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Download

PDBx/mmCIF format	5dyz.cif.gz	390.9 KB	Display	PDBx/mmCIF format
PDB format	pdb5dyz.ent.gz	319.9 KB	Display	PDB format
PDBx/mmJSON format	5dyz.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	5dyz_validation.pdf.gz	1.3 MB	Display	wwPDB validaton report
Full document	5dyz_full_validation.pdf.gz	1.3 MB	Display
Data in XML	5dyz_validation.xml.gz	40.4 KB	Display
Data in CIF	5dyz_validation.cif.gz	58.6 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/dy/5dyz ftp://data.pdbj.org/pub/pdb/validation_reports/dy/5dyz	HTTPS FTP

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Related structure data

Related structure data	5dypC 1jpzS S: Starting model for refinement C: citing same article (ref.)
Similar structure data	5dyp-d 5jq2-d 5e7y-d 6l1a-d 4dub-d 6jlv-d 4hgh-d 6k58-d 4dua-d 4duc-d 2bmh-d 6js8-d 1bu7-d 6k9s-d 6jvc-d 6l1b-d 5b2u-d 4hgf-d 1zo4-d 6jzs-d 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	#228 - Dec 2018 Directed Evolution of Enzymes similarity (14) #82 - Oct 2006 Cytochrome p450 similarity (2) #133 - Jan 2011 Nitric Oxide Synthase similarity (10)

Deposited unit

A: Bifunctional P-450/NADPH-P450 reductase

C: Bifunctional P-450/NADPH-P450 reductase

hetero molecules

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

1

Idetical with deposited unit
defined by software

Unit cell

Length a, b, c (Å)	61.213, 115.708, 143.278
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	19
Space group name H-M	P2₁2₁2₁

#1: Protein	Bifunctional P-450/NADPH-P450 reductase / Cytochrome P450(BM-3) / Cytochrome P450BM-3 Mass: 53591.078 Da / Num. of mol.: 2 / Mutation: D251G, Q307H Source method: isolated from a genetically manipulated source Source: (gene. exp.) Bacillus megaterium (bacteria) / Gene: cyp102A1, cyp102 / Production host: Escherichia coli (E. coli) References: UniProt: P14779, unspecific monooxygenase, NADPH-hemoprotein reductase
#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-140 / N-PALMITOYLGLYCINE / N-HEXADECANOYLGLYCINE Mass: 313.475 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₁₈H₃₅NO₃
#4: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 590 / Source method: isolated from a natural source / Formula: H₂O

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Experiment

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

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Sample preparation

Crystal	Density Matthews: 2.37 Å³/Da / Density % sol: 48.04 %
Crystal grow	Temperature: 277 K / Method: vapor diffusion, sitting drop Details: 14% PEG 3350, 100 mM cacodylic acid pH 5.5-6.8, 100-160 mM MgCl2

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Data collection

Diffraction

Mean temperature: 100 K

Diffraction source

Source:

SYNCHROTRON / Site:

ESRF

/ Beamline: ID29 / Wavelength: 0.9793 Å

Detector

Type: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Mar 15, 2014

Radiation

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

Radiation wavelength

Wavelength: 0.9793 Å / Relative weight: 1

Reflection

Resolution: 1.967→46.54 Å / Num. obs: 72820 / % possible obs: 99.6 % / Redundancy: 6.5 % / B_iso Wilson estimate: 31.3 Å² / CC_1/2: 0.998 / R_merge(I) obs: 0.093 / R_pim(I) all: 0.039 / Net I/σ(I): 12 / Num. measured all: 474709 / Scaling rejects: 4

Reflection shell

Diffraction-ID: 1 / Rejects: _

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Mean I/σ(I) obs	Num. measured all	Num. unique all	CC_1/2	R_pim(I) all	% possible all
1.967-2.01	6	0.864	2.1	25457	4220	0.67	0.376	94.7
9.64-46.54	5.7	0.039	33.4	4112	724	0.997	0.017	99

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Processing

Software

Name	Version	Classification
PHENIX	1.9_1692	refinement
XDS		data reduction
Aimless		data scaling
MOLREP		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 1JPZ

1jpz

Resolution: 1.967→46.538 Å / SU ML: 0.24 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 21.74 / Stereochemistry target values: ML

	R_factor	Num. reflection	% reflection	Selection details
R_free	0.219	3667	5.04 %	Random selection
R_work	0.179	-	-	-
obs	0.181	72738	99.54 %	-

Solvent computation

Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL

Displacement parameters

B_iso mean: 40 Å²

Refinement step

Cycle: LAST / Resolution: 1.967→46.538 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	7243	0	108	590	7941

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.01	7544
X-RAY DIFFRACTION	f_angle_d	1.309	10214
X-RAY DIFFRACTION	f_dihedral_angle_d	13.527	2850
X-RAY DIFFRACTION	f_chiral_restr	0.05	1086
X-RAY DIFFRACTION	f_plane_restr	0.006	1322

LS refinement shell

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Refine-ID	% reflection obs (%)
1.967-1.9929	0.3853	122	0.3245	2442	X-RAY DIFFRACTION	92
1.9929-2.0202	0.3143	136	0.2649	2607	X-RAY DIFFRACTION	100
2.0202-2.0491	0.2676	144	0.2425	2664	X-RAY DIFFRACTION	100
2.0491-2.0797	0.3101	149	0.2292	2607	X-RAY DIFFRACTION	100
2.0797-2.1122	0.3017	142	0.2276	2649	X-RAY DIFFRACTION	100
2.1122-2.1468	0.301	142	0.2183	2581	X-RAY DIFFRACTION	100
2.1468-2.1838	0.2855	140	0.2081	2646	X-RAY DIFFRACTION	100
2.1838-2.2235	0.2375	142	0.1935	2613	X-RAY DIFFRACTION	100
2.2235-2.2663	0.2343	158	0.1886	2641	X-RAY DIFFRACTION	100
2.2663-2.3125	0.2198	129	0.1866	2622	X-RAY DIFFRACTION	100
2.3125-2.3628	0.2384	138	0.1862	2701	X-RAY DIFFRACTION	100
2.3628-2.4178	0.2404	125	0.1901	2614	X-RAY DIFFRACTION	100
2.4178-2.4782	0.2515	152	0.1817	2658	X-RAY DIFFRACTION	100
2.4782-2.5452	0.2086	145	0.1778	2635	X-RAY DIFFRACTION	100
2.5452-2.6201	0.2299	148	0.1788	2636	X-RAY DIFFRACTION	100
2.6201-2.7047	0.2352	143	0.1817	2649	X-RAY DIFFRACTION	100
2.7047-2.8013	0.2256	113	0.1745	2683	X-RAY DIFFRACTION	100
2.8013-2.9135	0.2031	144	0.1855	2682	X-RAY DIFFRACTION	100
2.9135-3.0461	0.2427	147	0.191	2665	X-RAY DIFFRACTION	100
3.0461-3.2066	0.2512	145	0.1882	2656	X-RAY DIFFRACTION	100
3.2066-3.4075	0.2109	141	0.1753	2685	X-RAY DIFFRACTION	100
3.4075-3.6705	0.1977	138	0.1729	2671	X-RAY DIFFRACTION	100
3.6705-4.0397	0.2024	148	0.1515	2702	X-RAY DIFFRACTION	100
4.0397-4.6238	0.1814	145	0.1434	2728	X-RAY DIFFRACTION	100
4.6238-5.8237	0.1827	149	0.1657	2734	X-RAY DIFFRACTION	100
5.8237-46.5509	0.1925	142	0.1744	2900	X-RAY DIFFRACTION	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	4.6228	-0.9808	0.1334	3.5922	-0.1575	1.6142	0.1887	0.8584	0.7456	-0.3834	-0.0076	0.0196	-0.3796	-0.2139	-0.1884	0.3217	0.0569	0.1035	0.4642	0.1183	0.3436	-21.8051	-8.5486	-1.9368
2	5.0242	1.6533	-3.2441	2.1783	-1.6779	3.1152	-0.0742	-0.1121	-0.2856	-0.0797	-0.0665	-0.4504	0.0728	0.262	0.1097	0.1726	0.0218	-0.0294	0.2863	-0.094	0.211	-12.4049	-20.3006	21.5128
3	3.9436	1.4268	-1.7921	1.0505	-0.4244	2.6645	0.2621	0.0598	0.7858	0.0755	0.1429	0.0904	-0.492	-0.0464	-0.4128	0.2573	0.0599	0.0692	0.3329	-0.0149	0.3374	-25.4266	-12.9902	28.9345
4	9.1352	-4.826	1.2052	2.1671	-4.1315	9.1586	0.2025	-0.2011	1.4571	0.9312	-0.1739	-0.694	-1.708	0.165	0.0801	0.6234	-0.0639	0.0585	0.3685	-0.0828	0.564	-12.3437	-5.3235	30.4431
5	8.851	-1.2452	-4.4354	6.0076	2.6909	6.322	-0.0295	-0.9051	-0.1055	0.299	-0.0038	-0.3609	0.4262	1.0539	0.0355	0.2367	0.0917	-0.0537	0.5649	-0.0102	0.43	0.1817	-22.4135	27.7208
6	1.2156	0.376	-0.9353	1.0157	-0.0491	1.7164	0.0394	0.2182	-0.0013	-0.1223	-0.0064	-0.0707	0.0101	-0.0806	-0.0272	0.1427	0.044	0.0077	0.2677	-0.009	0.1986	-24.55	-25.3604	12.7395
7	5.1811	1.0795	-2.0282	4.5912	-2.3786	4.3881	0.2113	0.088	0.2216	0.1627	0.0106	0.2564	-0.2094	-0.2968	-0.2165	0.1376	0.0226	0.0142	0.209	-0.0605	0.138	-37.2434	-19.535	23.2317
8	2.9627	0.3824	-1.3086	1.851	1.5575	3.9104	0.0388	-0.5302	0.7594	0.5944	0.139	0.0209	-0.1214	0.2979	-0.1725	0.453	0.0703	-0.0662	0.2823	-0.118	0.4458	-40.9916	1.813	78.9387
9	3.9207	-1.1751	-1.5519	1.3676	1.4947	2.4236	-0.2129	0.0925	-0.151	0.3372	0.0329	0.1484	0.4479	-0.0145	0.1579	0.3061	0.0119	0.0246	0.1524	0.0422	0.1509	-40.7399	-17.6906	57.1409
10	2.5521	-1.7258	-0.9116	8.2074	6.0253	6.6066	0.153	0.3084	0.5088	-0.4315	-0.1863	0.0918	-0.7673	-0.0739	-0.0409	0.2974	-0.0108	0.0307	0.2664	0.0939	0.3111	-39.8119	-4.2159	52.3495
11	3.8031	2.1588	0.7455	2.1552	3.5918	6.1164	-0.4553	0.5872	0.9441	-1.1971	0.2663	1.0697	-1.2893	-0.2396	0.1106	0.4405	0.0407	-0.0509	0.3771	0.1421	0.4187	-48.4371	-3.0939	48.2962
12	7.4236	0.891	1.3145	5.1345	-0.5296	5.1977	-0.4004	0.5552	-0.2779	0.126	0.0709	0.3002	0.9584	-0.8051	0.4194	0.4508	-0.2114	0.1008	0.3364	-0.0598	0.3699	-57.7052	-24.6989	55.3584
13	5.0988	1.2008	2.3152	0.8019	0.6404	3.2266	-0.1652	0.1254	0.0012	0.2807	0.1154	-0.0713	0.1668	0.1577	0.0782	0.2885	0.0475	-0.0102	0.1427	-0.012	0.2427	-38.4286	-13.4589	59.2906
14	2.4053	-0.6536	-0.9825	1.7264	0.695	2.9175	-0.2087	-0.5227	-0.0829	0.6039	0.2094	-0.0948	0.746	0.679	0.0049	0.6977	0.2551	-0.1279	0.3274	-0.0081	0.316	-31.6637	-16.6295	74.5906
15	3.326	-1.6143	0.1855	2.5749	1.3664	1.9579	-0.3287	0.0123	-0.1618	0.686	0.0924	0.0466	1.0631	0.3451	0.1614	0.6179	0.1769	-0.0249	0.2245	0.0445	0.2859	-33.138	-20.9836	66.2957
16	4.4834	-2.643	-2.4767	5.7439	1.3258	3.0709	0.2481	-0.0568	0.401	-0.0186	-0.0322	-0.81	0.0798	0.7931	-0.1466	0.2558	0.0685	-0.1618	0.4221	0.0073	0.3429	-21.9509	-11.0787	57.7717

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	chain 'A' and (resid 2 through 72 )
2	X-RAY DIFFRACTION	2	chain 'A' and (resid 73 through 131 )
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 132 through 200 )
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 201 through 224 )
5	X-RAY DIFFRACTION	5	chain 'A' and (resid 225 through 250 )
6	X-RAY DIFFRACTION	6	chain 'A' and (resid 251 through 424 )
7	X-RAY DIFFRACTION	7	chain 'A' and (resid 425 through 456 )
8	X-RAY DIFFRACTION	8	chain 'C' and (resid 2 through 72 )
9	X-RAY DIFFRACTION	9	chain 'C' and (resid 73 through 158 )
10	X-RAY DIFFRACTION	10	chain 'C' and (resid 159 through 195 )
11	X-RAY DIFFRACTION	11	chain 'C' and (resid 196 through 224 )
12	X-RAY DIFFRACTION	12	chain 'C' and (resid 225 through 250 )
13	X-RAY DIFFRACTION	13	chain 'C' and (resid 251 through 282 )
14	X-RAY DIFFRACTION	14	chain 'C' and (resid 283 through 384 )
15	X-RAY DIFFRACTION	15	chain 'C' and (resid 385 through 424 )
16	X-RAY DIFFRACTION	16	chain 'C' and (resid 425 through 455 )

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