PDB-6iao: Structure of Cytochrome P450 BM3 M11 mutant in complex with DTT a...

ID or keywords:

Entry

Database: PDB / ID: 6iao

Title

Structure of Cytochrome P450 BM3 M11 mutant in complex with DTT at resolution 2.16A

Components

Bifunctional cytochrome P450/NADPH--P450 reductase

Keywords

OXIDOREDUCTASE /

Cytochrome P450 BM3 / mutant M11 / DTT

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 / FMN binding / iron ion binding /

heme binding / identical protein binding /

cytoplasm
Similarity search - Function

Biological species

Bacillus megaterium (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT /

molecular replacement / Resolution: 2.16 Å

Authors

Mirza, O. / Rafiq, M. / Frydenvang, K.

Citation

Journal: Plos One / Year: 2019
Title: Structural analysis of Cytochrome P450 BM3 mutant M11 in complex with dithiothreitol.
Authors: Frydenvang, K. / Verkade-Vreeker, M.C.A. / Dohmen, F. / Commandeur, J.N.M. / Rafiq, M. / Mirza, O. / Jorgensen, F.S. / Geerke, D.P.

History

Deposition	Nov 27, 2018	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Jun 5, 2019	Provider: repository / Type: Initial release
Revision 1.1	Jan 24, 2024	Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_conn Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_conn.conn_type_id / _struct_conn.id / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_conn.ptnr2_label_seq_id

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	6iao.cif.gz	758.4 KB	Display	PDBx/mmCIF format
PDB format	pdb6iao.ent.gz	626 KB	Display	PDB format
PDBx/mmJSON format	6iao.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/ia/6iao ftp://data.pdbj.org/pub/pdb/validation_reports/ia/6iao	HTTPS FTP

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Related structure data

Related structure data	3dgiS S: Starting model for refinement
Similar structure data	3n9z-d 3na0-d 6std-2 6s3h-d 5w9c-d 3na1-d 3bt9-d 1hnc-d 2gby-d 6ns7-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	#228 - Dec 2018 Directed Evolution of Enzymes similarity (14) #133 - Jan 2011 Nitric Oxide Synthase similarity (10) #82 - Oct 2006 Cytochrome p450 similarity (1) #92 - Aug 2007 Anabolic Steroids similarity (1) #127 - Jul 2010 Crystallins similarity (1) #174 - Jun 2014 GFP-like Proteins similarity (1) #89 - May 2007 Aconitase and Iron Regulatory Protein 1 similarity (1)

Deposited unit

A: Bifunctional cytochrome P450/NADPH--P450 reductase

B: Bifunctional cytochrome P450/NADPH--P450 reductase

C: Bifunctional cytochrome P450/NADPH--P450 reductase

D: Bifunctional cytochrome P450/NADPH--P450 reductase

hetero molecules

234 kDa, 4 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: Bifunctional cytochrome P450/NADPH--P450 reductase

B: Bifunctional cytochrome P450/NADPH--P450 reductase

hetero molecules

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

2

C: Bifunctional cytochrome P450/NADPH--P450 reductase

D: Bifunctional cytochrome P450/NADPH--P450 reductase

hetero molecules

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

Unit cell

Length a, b, c (Å)	377.933, 59.937, 95.456
Angle α, β, γ (deg.)	90.000, 95.740, 90.000
Int Tables number	5
Space group name H-M	C121

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Protein , 1 types, 4 molecules ABCD

#1: Protein	Bifunctional cytochrome P450/NADPH--P450 reductase / Cytochrome P450(BM-3) / Cytochrome P450BM-3 / Fatty acid monooxygenase / Flavocytochrome P450 BM3 Mass: 57319.395 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Bacillus megaterium (bacteria) / Gene: cyp102A1, cyp102, BG04_163 / Plasmid: PET-28A(+) / Production host: Escherichia coli BL21 (bacteria) References: UniProt: P14779, unspecific monooxygenase, NADPH-hemoprotein reductase

#1: Protein

Bifunctional cytochrome P450/NADPH--P450 reductase / Cytochrome P450(BM-3) / Cytochrome P450BM-3 / Fatty acid monooxygenase / Flavocytochrome P450 BM3

Mass: 57319.395 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.)

Bacillus megaterium (bacteria) / Gene: cyp102A1, cyp102, BG04_163 / Plasmid: PET-28A(+) / Production host:

Escherichia coli BL21 (bacteria)
References: UniProt: P14779,

unspecific monooxygenase,

NADPH-hemoprotein reductase

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Non-polymers , 6 types, 1280 molecules

#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-DTT / 2,3-DIHYDROXY-1,4-DITHIOBUTANE / 1,4-DITHIOTHREITOL / Dithiothreitol Mass: 154.251 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C₄H₁₀O₂S₂
#4: Chemical	ChemComp-CL / CHLORIDE ION / Chloride Mass: 35.453 Da / Num. of mol.: 20 / Source method: obtained synthetically / Formula: Cl
#5: Chemical	ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL / Glycerol Mass: 92.094 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C₃H₈O₃
#6: Chemical	ChemComp-PEG / DI(HYDROXYETHYL)ETHER / Diethylene glycol Mass: 106.120 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₄H₁₀O₃
#7: Water	ChemComp-HOH / water / Water Mass: 18.015 Da / Num. of mol.: 1245 / 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: 2.6 Å³/Da / Density % sol: 52.67 %
Crystal grow	Temperature: 279 K / Method: vapor diffusion, hanging drop / pH: 8 Details: 18% PEG4000, 0.25 M Magnesium chloride, 0.1 M TRIS pH 8.0, 10 mM DTT

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

MAX II

/ Beamline: I911-3 / Wavelength: 1 Å

Detector

Type: MARMOSAIC 225 mm CCD / Detector: CCD / Date: Oct 6, 2015

Radiation

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

Radiation wavelength

Wavelength: 1 Å / Relative weight: 1

Reflection

Resolution: 2.16→188.019 Å / Num. obs: 114389 / % possible obs: 99.6 % / Redundancy: 5.1 % / B_iso Wilson estimate: 20.12 Å² / R_pim(I) all: 0.067 / R_rim(I) all: 0.155 / R_sym value: 0.139 / Net I/av σ(I): 4.2 / Net I/σ(I): 13.5

Reflection shell

Diffraction-ID: 1

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Mean I/σ(I) obs	Num. unique obs	R_pim(I) all	R_rim(I) all	R_sym value	% possible all
2.16-2.28	3.9	0.727	0.9	16254	0.404	0.837	0.727	97.6
2.28-2.41	5.2	0.507	1.4	15721	0.241	0.563	0.507	100
2.41-2.58	5.3	0.367	1.8	14854	0.172	0.406	0.367	100
2.58-2.79	5.3	0.271	2.4	13804	0.127	0.299	0.271	100
2.79-3.05	5.3	0.171	3.7	12728	0.08	0.19	0.171	100
3.05-3.42	5.3	0.109	5.9	11573	0.051	0.12	0.109	100
3.42-3.94	5.3	0.07	5.6	10189	0.033	0.077	0.07	100
3.94-4.83	5.3	0.046	14.4	8668	0.021	0.05	0.046	100
4.83-6.83	5.3	0.049	13.4	6773	0.023	0.054	0.049	100
6.83-47.888	5	0.032	13.4	3825	0.016	0.036	0.032	99.7

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Phasing

Phasing

Method:

molecular replacement

Phasing MR

Model details: Phaser MODE: MR_AUTO

	Highest resolution	Lowest resolution
Rotation	4.83 Å	47.88 Å
Translation	4.83 Å	47.88 Å

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Processing

Software

Name	Version	Classification
SCALA	3.3.20	data scaling
PHASER	2.5.1	phasing
PHENIX		refinement
PDB_EXTRACT	3.24	data extraction
XDS		data reduction

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 3DGI

3dgi

Resolution: 2.16→47.177 Å / SU ML: 0.23 / Cross valid method: THROUGHOUT / σ(F): 1.37 / Phase error: 18.82

	R_factor	Num. reflection	% reflection
R_free	0.2021	5743	5.02 %
R_work	0.1589	-	-
obs	0.1611	114358	99.62 %

Solvent computation

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

Displacement parameters

B_iso max: 97.79 Å² / B_iso mean: 26.7173 Å² / B_iso min: 3.68 Å²

Refinement step

Cycle: final / Resolution: 2.16→47.177 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	14450	0	270	1256	15976
B_iso mean	-	-	29.52	27.25	-
Num. residues	-	-	-	-	1807

LS refinement shell

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

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Num. reflection all	% reflection obs (%)
2.16-2.1846	0.2789	176	0.2476	3280	3456	92
2.1846-2.2103	0.2761	196	0.2315	3542	3738	98
2.2103-2.2372	0.3201	199	0.2323	3559	3758	100
2.2372-2.2655	0.284	181	0.2448	3610	3791	100
2.2655-2.2953	0.2686	191	0.1961	3628	3819	100
2.2953-2.3268	0.2447	200	0.1846	3612	3812	100
2.3268-2.36	0.2246	196	0.1759	3577	3773	100
2.36-2.3953	0.2355	184	0.1747	3659	3843	100
2.3953-2.4327	0.2162	191	0.1758	3569	3760	100
2.4327-2.4726	0.2449	185	0.1711	3604	3789	100
2.4726-2.5152	0.2201	211	0.1629	3639	3850	100
2.5152-2.5609	0.2096	178	0.1704	3596	3774	100
2.5609-2.6102	0.2525	169	0.1744	3675	3844	100
2.6102-2.6635	0.2377	180	0.1737	3568	3748	100
2.6635-2.7214	0.2307	191	0.1691	3648	3839	100
2.7214-2.7847	0.208	176	0.1618	3608	3784	100
2.7847-2.8543	0.2104	179	0.1593	3657	3836	100
2.8543-2.9315	0.2072	207	0.1631	3576	3783	100
2.9315-3.0177	0.2033	197	0.1623	3648	3845	100
3.0177-3.1151	0.1956	192	0.1749	3623	3815	100
3.1151-3.2264	0.2236	207	0.1616	3618	3825	100
3.2264-3.3555	0.2069	177	0.1549	3655	3832	100
3.3555-3.5082	0.1871	176	0.1453	3666	3842	100
3.5082-3.6931	0.193	180	0.1396	3643	3823	100
3.6931-3.9244	0.166	195	0.1319	3640	3835	100
3.9244-4.2272	0.1498	202	0.12	3661	3863	100
4.2272-4.6523	0.1336	198	0.1104	3677	3875	100
4.6523-5.3247	0.1503	211	0.1203	3665	3876	100
5.3247-6.7055	0.198	191	0.1533	3724	3915	100
6.7055-47.188	0.1836	227	0.1635	3788	4015	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	1.8662	0.5047	-0.2485	1.9913	-0.5922	1.9947	-0.0162	-0.0414	-0.3924	-0.0212	-0.0218	0.0756	0.3447	-0.16	0.0069	0.1629	-0.0039	0.0106	0.1167	-0.0056	0.1692	56.7841	-14.3089	-16.9209
2	1.9073	-0.4241	0.9358	0.795	-0.4336	1.6072	0.0427	0.1884	-0.1188	-0.1049	-0.0207	-0.0773	0.1054	0.2333	-0.0175	0.1506	0.0253	0.043	0.1508	-0.0157	0.1074	72.5687	-5.5964	-27.5946
3	3.8573	-0.8182	2.3098	1.8124	-0.6707	4.6537	-0.1553	-0.0621	0.2393	0.0019	0.0833	-0.0911	-0.2469	-0.2377	0.0649	0.1147	-0.006	0.0524	0.1039	0.0076	0.09	61.7691	9.6377	-25.0352
4	0.6979	-0.1734	-0.6751	1.2583	-0.2659	2.3844	0.0692	-0.0706	-0.0343	-0.0013	-0.022	0.0791	-0.0318	-0.054	-0.0384	0.089	0.0059	-0.0061	0.1617	-0.0199	0.2166	-6.4153	25.1775	82.657
5	2.3091	-1.376	-0.069	3.3977	-0.1933	0.7048	-0.0781	-0.1276	0.1071	0.1802	0.1206	0.0463	-0.1306	0.0139	-0.0163	0.1375	0.0069	0.0081	0.1349	0.0067	0.1338	20.3399	17.4476	70.837
6	2.4184	-0.2049	-0.1183	3.7072	-0.395	1.2597	0.06	0.1687	0.3964	0.0037	-0.0407	-0.372	-0.2394	0.1699	0.0264	0.1473	0.0254	-0.0038	0.1745	-0.0027	0.2013	22.6365	24.3313	66.3845
7	2.0621	-1.1776	-0.5317	1.1106	0.3474	0.5053	0.0294	0.1288	-0.4292	-0.052	-0.1311	0.3386	0.0956	-0.1194	0.0795	0.1473	-0.0186	-0.0292	0.1427	-0.0335	0.2707	5.0419	8.9327	69.1318
8	2.0259	0.506	0.4778	0.8969	-0.0467	0.6398	-0.1155	0.1487	0.2106	-0.0932	0.0913	0.2385	-0.0307	-0.0698	-0.0032	0.1631	-0.0264	-0.0053	0.1472	0.0163	0.1709	16.108	44.8314	114.4876
9	2.9488	-0.0579	-0.0128	2.3977	0.3471	2.0263	0.0607	0.1199	-0.2863	-0.011	0.0835	0.1592	0.2219	-0.4412	-0.0687	0.1932	-0.0365	-0.0375	0.27	0.0017	0.2554	16.6594	28.9726	120.9105
10	1.5942	0.5551	0.498	1.1029	-0.0042	0.9098	-0.1072	0.0496	0.3684	-0.0476	0.079	0.2919	-0.1928	-0.0571	0.0509	0.165	0.0014	-0.0085	0.1276	0.0159	0.2382	19.2611	55.1719	118.0613
11	3.8354	-0.2272	2.3995	1.1325	0.5347	5.1088	0.1568	0.2314	-0.0482	-0.0418	-0.0296	0.0985	0.174	0.073	-0.1125	0.1787	-0.0461	0.009	0.0559	0.0319	0.1494	34.8119	49.694	112.5764
12	2.087	1.2677	0.7072	2.3251	1.1699	2.221	0.0662	-0.0648	-0.173	0.0679	-0.0241	-0.2235	0.2321	0.1177	-0.0287	0.1378	0.0181	-0.0001	0.1059	0.0388	0.1351	81.0027	3.2958	27.3385
13	3.2785	-0.7003	-1.2445	0.7338	0.2702	1.1192	0.0262	-0.1896	0.1581	0.0347	0.0227	-0.1674	-0.0872	0.163	-0.0492	0.14	-0.0232	-0.0251	0.0854	0.0103	0.0894	74.444	30.8331	28.6554
14	0.2595	-0.5855	-0.2944	0.9522	0.6453	0.9021	-0.0233	0.1303	-0.1218	-0.0798	0.0119	0.017	0.0234	-0.0511	0.0273	0.1526	-0.0195	-0.0164	0.1816	-0.0223	0.1771	63.2055	26.019	19.7195
15	1.5585	0.5088	-0.9204	2.1592	-1.0391	1.8702	0.0292	-0.0033	0.1491	-0.0647	-0.0789	-0.2822	-0.1015	0.1585	0.0198	0.125	-0.0226	-0.0054	0.13	-0.0063	0.1353	74.6244	33.0172	26.3115
16	1.4974	-0.769	-0.6444	1.1568	0.2376	0.8671	-0.114	-0.3114	-0.0069	0.2421	0.0928	-0.1008	0.0269	0.1334	0.0217	0.1759	-0.0131	-0.0378	0.2033	0.0125	0.0991	73.1617	18.7714	42.2174
17	2.8465	-1.8913	-1.6372	3.0576	2.1653	2.9427	0.0666	-0.0253	-0.1037	0.0598	-0.0692	0.1666	-0.0563	-0.1732	0.0106	0.1308	-0.0197	-0.004	0.1456	0.0563	0.0983	55.3758	18.8782	36.8105
18	1.893	-0.7565	0.547	1.974	0.3086	3.7043	-0.1314	-0.4637	0.1562	0.2505	0.244	-0.2537	-0.3229	0.3314	-0.0509	0.1694	-0.0096	0.0036	0.2978	-0.076	0.1683	82.9855	3.5588	-4.9135
19	1.7666	-0.0247	0.3988	0.5638	-0.1073	0.8602	0.0219	-0.0665	-0.1925	0.0362	0.0227	0.0207	0.1061	0.0103	-0.0368	0.1411	0.0246	0.0245	0.0923	-0.0053	0.1102	62.9655	-8.4225	-17.0571
20	3.6435	1.443	2.6764	1.1205	1.6404	3.3141	0.1233	-0.4067	0.0237	0.2295	-0.1352	0.0744	0.1191	-0.2293	0.0759	0.1739	0.0113	0.0192	0.1654	0.0155	0.1488	56.9805	-3.4015	-1.6912

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	chain 'B' and (resid 225 through 282 )	B	225 - 282
2	X-RAY DIFFRACTION	2	chain 'B' and (resid 283 through 424 )	B	283 - 424
3	X-RAY DIFFRACTION	3	chain 'B' and (resid 425 through 456 )	B	425 - 456
4	X-RAY DIFFRACTION	4	chain 'C' and (resid -5 through 72 )	C	-5 - 72
5	X-RAY DIFFRACTION	5	chain 'C' and (resid 73 through 205 )	C	73 - 205
6	X-RAY DIFFRACTION	6	chain 'C' and (resid 206 through 281 )	C	206 - 281
7	X-RAY DIFFRACTION	7	chain 'C' and (resid 282 through 457 )	C	282 - 457
8	X-RAY DIFFRACTION	8	chain 'D' and (resid 4 through 171 )	D	4 - 171
9	X-RAY DIFFRACTION	9	chain 'D' and (resid 172 through 250 )	D	172 - 250
10	X-RAY DIFFRACTION	10	chain 'D' and (resid 251 through 424 )	D	251 - 424
11	X-RAY DIFFRACTION	11	chain 'D' and (resid 425 through 458 )	D	425 - 458
12	X-RAY DIFFRACTION	12	chain 'A' and (resid 3 through 72 )	A	3 - 72
13	X-RAY DIFFRACTION	13	chain 'A' and (resid 73 through 132 )	A	73 - 132
14	X-RAY DIFFRACTION	14	chain 'A' and (resid 133 through 225 )	A	133 - 225
15	X-RAY DIFFRACTION	15	chain 'A' and (resid 226 through 281 )	A	226 - 281
16	X-RAY DIFFRACTION	16	chain 'A' and (resid 282 through 424 )	A	282 - 424
17	X-RAY DIFFRACTION	17	chain 'A' and (resid 425 through 457 )	A	425 - 457
18	X-RAY DIFFRACTION	18	chain 'B' and (resid 5 through 36 )	B	5 - 36
19	X-RAY DIFFRACTION	19	chain 'B' and (resid 37 through 171 )	B	37 - 171
20	X-RAY DIFFRACTION	20	chain 'B' and (resid 172 through 224 )	B	172 - 224

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