PDB-8dh0: T7 RNA polymerase elongation complex with unnatural base dDs

ID or keywords:

Entry

Database: PDB / ID: 8dh0

Title

T7 RNA polymerase elongation complex with unnatural base dDs

Components

Non-template strand DNA
RNA
T7 RNA polymerase
Template strand DNA

Keywords

Transferase/DNA/RNA / T7 RNA polymerase / elongation complex / unnatural base / Ds-Pa / synthetic DNA / transcription / Transferase-DNA-RNA complex

Function / homology

Function and homology information

DNA-templated viral transcription / DNA-directed RNA polymerase complex / : / : / : / : / : / : / DNA-directed RNA polymerase / DNA binding
Similarity search - Function

Biological species

Escherichia phage T7 (virus)
synthetic construct (others)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.9 Å

Model details

T7 RNA polymerase, elongation complex, unnatural base pair, synthetic DNA

Authors

Oh, J. / Wang, D.

Funding support

United States, 1items

Citation

Journal: Nat Commun / Year: 2023
Title: Structural basis of transcription recognition of a hydrophobic unnatural base pair by T7 RNA polymerase.
Authors: Oh, J. / Kimoto, M. / Xu, H. / Chong, J. / Hirao, I. / Wang, D.

History

Deposition	Jun 24, 2022	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Feb 1, 2023	Provider: repository / Type: Initial release
Revision 1.1	Oct 25, 2023	Group: Data collection / Refinement description Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	8dh0.cif.gz	1.3 MB	Display	PDBx/mmCIF format
PDB format	pdb8dh0.ent.gz	1.1 MB	Display	PDB format
PDBx/mmJSON format	8dh0.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	8dh0_validation.pdf.gz	563 KB	Display	wwPDB validaton report
Full document	8dh0_full_validation.pdf.gz	628.6 KB	Display
Data in XML	8dh0_validation.xml.gz	109.9 KB	Display
Data in CIF	8dh0_validation.cif.gz	149.6 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/dh/8dh0 ftp://data.pdbj.org/pub/pdb/validation_reports/dh/8dh0	HTTPS FTP

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Related structure data

Related structure data	8dh1C 8dh2C 8dh3C 8dh4C 8dh5C 1h38S S: Starting model for refinement C: citing same article (ref.)
Similar structure data	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
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PDB pages

PDBj /

wwPDB /

NCBI

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Deposited unit

A: Template strand DNA

B: T7 RNA polymerase

C: RNA

E: Template strand DNA

F: T7 RNA polymerase

G: RNA

H: Non-template strand DNA

I: Template strand DNA

J: T7 RNA polymerase

K: RNA

M: Template strand DNA

N: T7 RNA polymerase

O: RNA

P: Non-template strand DNA

hetero molecules

440 kDa, 14 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

Idetical with deposited unit
defined by author

Unit cell

Length a, b, c (Å)	78.707, 86.235, 201.220
Angle α, β, γ (deg.)	89.910, 85.230, 69.500
Int Tables number	1
Space group name H-M	P1

#1: DNA chain	Template strand DNA Mass: 5616.727 Da / Num. of mol.: 4 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#2: Protein	T7 RNA polymerase / DNA-directed RNA polymerase Mass: 98984.227 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Escherichia phage T7 (virus) / References: UniProt: P00573, DNA-directed RNA polymerase
#3: RNA chain	RNA Mass: 3851.360 Da / Num. of mol.: 4 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#4: DNA chain	Non-template strand DNA Mass: 2666.761 Da / Num. of mol.: 2 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#5: Chemical	ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL Mass: 92.094 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C₃H₈O₃
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: 2.98 Å³/Da / Density % sol: 58.67 % / Mosaicity: 0.25 °
Crystal grow	Temperature: 298 K / Method: vapor diffusion, hanging drop / pH: 8.1 Details: 10% PEG 8000, 8% glycerol, 5 mM B-mercaptoethanol, 100 mM Tris pH 8.1

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

SSRL

/ Beamline: BL12-1 / Wavelength: 1 Å

Detector

Type: DECTRIS EIGER X 16M / Detector: PIXEL / Date: Jul 16, 2021

Radiation

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

Radiation wavelength

Wavelength: 1 Å / Relative weight: 1

Reflection

Resolution: 2.66→42.03 Å / Num. obs: 131648 / % possible obs: 89.7 % / Redundancy: 3.9 % / B_iso Wilson estimate: 84.98 Å² / CC_1/2: 0.997 / R_merge(I) obs: 0.099 / R_pim(I) all: 0.058 / R_rim(I) all: 0.115 / Net I/σ(I): 6.9

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
2.66-2.7	3.7	5.785	24203	6456	0.234	3.403	6.716	0.3	88.9
14.57-39.64	3.8	0.035	3232	851	0.996	0.021	0.041	25.6	94.8

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Processing

Software

Name	Version	Classification
xia2		data scaling
PHENIX	1.19	refinement
PDB_EXTRACT	3.27	data extraction
DIALS		data reduction
PHENIX	1.19	phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 1H38

1h38

Resolution: 2.9→42.03 Å / SU ML: 0.48 / Cross valid method: THROUGHOUT / σ(F): 1.96 / Phase error: 26.35 / Stereochemistry target values: ML

	R_factor	Num. reflection	% reflection
R_free	0.261	1783	1.68 %
R_work	0.2207	104268	-
obs	0.2213	106051	96.91 %

Solvent computation

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

Displacement parameters

B_iso max: 279.82 Å² / B_iso mean: 101.1379 Å² / B_iso min: 29.85 Å²

Refinement step

Cycle: final / Resolution: 2.9→42.03 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	24142	2035	24	0	26201
B_iso mean	-	-	84.19	-	-
Num. residues	-	-	-	-	3311

LS refinement shell

Refine-ID: X-RAY DIFFRACTION / R_factor R_free error: 0 / Total num. of bins used: 13

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Num. reflection all	% reflection obs (%)
2.9-2.98	0.4449	123	0.4344	7102	7225	86
2.98-3.07	0.354	134	0.3528	7868	8002	95
3.07-3.16	0.3705	132	0.3057	7797	7929	94
3.16-3.28	0.3031	140	0.2649	8205	8345	99
3.28-3.41	0.2788	141	0.2606	8172	8313	99
3.41-3.56	0.2707	140	0.2516	8206	8346	99
3.56-3.75	0.2598	140	0.2188	8247	8387	99
3.75-3.99	0.2685	139	0.1971	8041	8180	98
3.99-4.29	0.2353	137	0.19	8077	8214	97
4.29-4.73	0.2279	140	0.1778	8214	8354	99
4.73-5.41	0.238	140	0.1961	8213	8353	99
5.41-6.81	0.2642	138	0.2315	7991	8129	97
6.81-42.03	0.2375	139	0.1991	8135	8274	98

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	1.6917	0.2739	0.4563	1.9224	0.2908	0.9524	0.0371	-0.6959	-0.6513	0.2961	-0.0107	-0.2511	0.3305	0.0294	-0.0181	0.6331	-0.0319	0.1233	0.8826	0.1556	0.8028	33.3011	57.5039	8.5994
2	2.2326	0.0547	-0.0524	1.3599	-0.1929	0.7635	-0.061	-0.521	0.1516	-0.0423	-0.1106	-0.0114	-0.1185	-0.0083	0.1663	0.4726	0.0274	-0.0011	0.4735	-0.1412	0.4499	36.825	82.3827	-6.2087
3	7.1538	-4.253	0.9234	2.8309	-1.4934	4.9306	0.2762	-1.1498	-0.4902	-0.7063	-0.388	-0.9275	1.1639	1.9367	0.2279	0.8098	0.2327	0.0689	1.1585	0.0363	0.7317	37.5564	64.3721	-2.393
4	4.1245	-0.9178	-0.1661	4.6344	-0.1539	0.0189	-0.6808	-0.1148	0.2278	0.6434	0.4365	-0.0371	0.307	-0.2145	0.2389	3.0835	-0.2174	0.3671	2.1732	-0.2095	2.0706	64.0783	76.6748	-0.4559
5	3.6845	-0.1203	0.2529	3.4127	0.2851	3.5508	0.3988	-0.438	1.2745	-0.3003	0.1841	1.8727	0.1993	-0.7633	-0.4945	0.8309	-0.2297	-0.1693	0.7542	0.1602	1.1479	1.4872	23.1185	-92.8692
6	5.9922	-1.8623	1.0661	1.2125	-0.7732	1.4274	-0.2067	-0.5505	1.2504	0.2136	0.148	-0.0949	-0.2218	0.1286	0.1979	0.7215	-0.1846	0.0567	0.6705	-0.1551	1.0255	28.4367	37.3861	-96.1339
7	2.75	0.5123	-0.5963	0.4572	0.0245	1.8159	0.0491	0.0343	1.334	0.167	0.0947	0.4304	-0.5503	-0.6376	-0.1622	0.8277	0.1649	0.0292	0.7164	-0.05	1.5174	-9.192	47.5775	-96.0641
8	3.0114	-0.2208	-0.041	2.3851	0.1543	3.179	0.2671	-0.0493	-0.1878	-0.3436	-0.1438	0.0857	0.4475	0.1326	-0.123	0.8039	-0.0387	-0.0809	0.5709	-0.0489	0.4028	22.0934	13.4003	-100.4699
9	3.6745	-0.2656	-0.0013	1.9893	0.2906	1.9605	0.2983	-0.6867	-0.4288	-0.247	-0.1077	0.6483	0.4341	-0.5275	-0.1801	0.9568	-0.2055	-0.1852	0.8094	0.1097	0.7193	1.7095	6.89	-85.1404
10	6.7176	-4.5041	2.1492	5.1678	-3.9696	3.6503	0.7602	-0.4766	0.247	-0.5381	-0.7016	1.7153	0.6796	-0.8942	-0.0209	1.0911	0.0071	-0.1459	0.6718	-0.2415	1.1264	4.6665	22.3826	-98.4087
11	6.6222	-2.5758	-0.8903	5.7719	0.4119	3.3759	-0.4804	0.1571	-1.0409	0.5281	0.0701	1.9042	-0.2758	-0.4099	0.2361	1.1698	0.0257	0.0535	0.9991	-0.1424	1.0308	4.8585	11.2425	53.0713
12	3.1395	0.6969	-0.6149	1.5546	-1.0524	2.2925	0.0811	0.6286	-0.2819	-0.0662	0.4802	0.4852	0.5782	-0.857	-0.4615	1.1988	-0.0219	-0.1955	1.1465	0.0987	0.9822	4.6268	-12.921	57.1876
13	2.1919	0.0686	1.1117	0.8381	0.3046	2.2497	0.0351	0.3727	0.206	-0.0217	-0.1801	0.091	-0.2651	0.2871	0.1353	0.9357	0.0322	-0.0797	0.8927	-0.0584	0.4976	22.7674	17.3937	49.0679
14	5.541	-0.1107	-1.955	6.9003	-2.4815	5.0121	-0.095	0.0125	-0.0367	0.1013	0.0322	2.2385	-0.4558	-0.4211	0.0447	1.1769	0.1379	-0.018	1.419	-0.3199	0.98	13.2488	8.4392	58.4707
15	3.7352	1.2467	-1.1294	5.4803	-3.2101	2.0228	-0.3868	0.3788	0.1206	0.3111	-0.4357	0.2832	0.9374	-0.7948	0.7422	3.1064	0.1037	0.5277	2.3139	-0.4773	2.158	-7.1932	25.4534	44.8782
16	1.8321	1.473	0.327	1.2476	0.1581	3.5881	0.1912	0.5747	0.4037	-0.4005	-0.3756	-0.6866	0.3783	1.3496	0.0931	0.476	0.0959	0.1031	0.8274	0.1731	0.7694	32.0918	37.5229	-44.6243
17	1.3646	0.2655	0.0953	0.0319	-0.124	1.1766	0.1312	1.4534	0.8152	-0.0928	-0.1584	-0.1415	-0.0989	0.411	-0.0028	0.5321	0.0623	0.1132	1.1158	0.4806	0.9971	35.4632	54.4399	-57.3216
18	1.9013	-0.2606	0.2002	1.0544	-0.1874	1.183	0.087	0.2417	-0.0094	0.0329	-0.0866	-0.0893	0.2145	0.0593	0.0144	0.3648	-0.0546	0.0219	0.3478	0.0218	0.4194	18.1883	31.0598	-35.3973
19	6.7688	3.9646	0.3378	6.2413	-0.3598	1.7533	0.3532	1.2137	0.2887	-0.4893	-0.2617	-0.9898	0.2262	1.1311	-0.1017	0.4066	0.0196	0.0718	0.7995	0.1285	0.7026	27.7672	43.3141	-40.1877
20	1.4505	-1.935	-0.4187	2.7457	0.8915	2.371	0.0901	-0.7375	-0.6444	-0.2358	-0.2822	-1.0112	0.2052	1.6214	0.2061	0.7574	0.0843	0.1206	1.3411	0.1646	1.0438	48.7091	67.3952	0.6892

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	chain 'F' and (resid 1 through 407 )	F	1 - 407
2	X-RAY DIFFRACTION	2	chain 'F' and (resid 408 through 883 )	F	408 - 883
3	X-RAY DIFFRACTION	3	chain 'G' and (resid 1 through 8 )	G	1 - 8
4	X-RAY DIFFRACTION	4	chain 'H' and (resid 2 through 7 )	H	2 - 7
5	X-RAY DIFFRACTION	5	chain 'I' and (resid 7 through 18 )	I	7 - 18
6	X-RAY DIFFRACTION	6	chain 'J' and (resid 2 through 60 )	J	2 - 60
7	X-RAY DIFFRACTION	7	chain 'J' and (resid 61 through 263 )	J	61 - 263
8	X-RAY DIFFRACTION	8	chain 'J' and (resid 264 through 527 )	J	264 - 527
9	X-RAY DIFFRACTION	9	chain 'J' and (resid 528 through 883 )	J	528 - 883
10	X-RAY DIFFRACTION	10	chain 'K' and (resid 1 through 8 )	K	1 - 8
11	X-RAY DIFFRACTION	11	chain 'M' and (resid 5 through 18 )	M	5 - 18
12	X-RAY DIFFRACTION	12	chain 'N' and (resid 1 through 264 )	N	1 - 264
13	X-RAY DIFFRACTION	13	chain 'N' and (resid 265 through 883 )	N	265 - 883
14	X-RAY DIFFRACTION	14	chain 'O' and (resid 1 through 8 )	O	1 - 8
15	X-RAY DIFFRACTION	15	chain 'P' and (resid 2 through 6 )	P	2 - 6
16	X-RAY DIFFRACTION	16	chain 'A' and (resid 6 through 18 )	A	6 - 18
17	X-RAY DIFFRACTION	17	chain 'B' and (resid 1 through 313 )	B	1 - 313
18	X-RAY DIFFRACTION	18	chain 'B' and (resid 314 through 883 )	B	314 - 883
19	X-RAY DIFFRACTION	19	chain 'C' and (resid 1 through 8 )	C	1 - 8
20	X-RAY DIFFRACTION	20	chain 'E' and (resid 4 through 18 )	E	4 - 18

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