PDB-8dh5: T7 RNA polymerase elongation complex with unnatural base dPa-ATP ...

ID or keywords:

Entry

Database: PDB / ID: 8dh5

Title

T7 RNA polymerase elongation complex with unnatural base dPa-ATP mismatch

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-directed RNA polymerase activity / DNA-templated transcription / DNA binding
Similarity search - Function

Biological species

Escherichia phage T7 (virus)
synthetic construct (others)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.85 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	8dh5.cif.gz	1.4 MB	Display	PDBx/mmCIF format
PDB format	pdb8dh5.ent.gz	1.2 MB	Display	PDB format
PDBx/mmJSON format	8dh5.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	8dh5_validation.pdf.gz	1.7 MB	Display	wwPDB validaton report
Full document	8dh5_full_validation.pdf.gz	1.8 MB	Display
Data in XML	8dh5_validation.xml.gz	113 KB	Display
Data in CIF	8dh5_validation.cif.gz	153.5 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/dh/8dh5 ftp://data.pdbj.org/pub/pdb/validation_reports/dh/8dh5	HTTPS FTP

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Related structure data

Related structure data	8dh0SC 8dh1C 8dh2C 8dh3C 8dh4C 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

B: T7 RNA polymerase

A: Template strand DNA

C: RNA

D: Non-template strand DNA

E: T7 RNA polymerase

F: Template strand DNA

G: RNA

H: Non-template strand DNA

I: T7 RNA polymerase

J: Template strand DNA

K: RNA

L: Non-template strand DNA

M: T7 RNA polymerase

N: Template strand DNA

O: RNA

P: Non-template strand DNA

hetero molecules

446 kDa, 16 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

B: T7 RNA polymerase

A: Template strand DNA

C: RNA

D: Non-template strand DNA

hetero molecules

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

2

E: T7 RNA polymerase

F: Template strand DNA

G: RNA

H: Non-template strand DNA

hetero molecules

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

3

I: T7 RNA polymerase

J: Template strand DNA

K: RNA

L: Non-template strand DNA

hetero molecules

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

4

M: T7 RNA polymerase

N: Template strand DNA

O: RNA

P: Non-template strand DNA

hetero molecules

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

Unit cell

Length a, b, c (Å)	78.544, 86.305, 201.199
Angle α, β, γ (deg.)	89.790, 85.390, 69.490
Int Tables number	1
Space group name H-M	P1

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DNA chain , 2 types, 8 molecules AFJNDHLP

#2: DNA chain	Template strand DNA Mass: 5510.578 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.: 4 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)

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Protein / RNA chain , 2 types, 8 molecules BEIMCGKO

#1: 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)

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Non-polymers , 2 types, 6 molecules

#5: Chemical	ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE Mass: 507.181 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C₁₀H₁₆N₅O₁₃P₃ / Feature type: SUBJECT OF INVESTIGATION / Comment: ATP, energy-carrying molecule*YM
#6: Chemical	ChemComp-MG / MAGNESIUM ION Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg

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Details

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: 2.87 Å³/Da / Density % sol: 57.09 % / Mosaicity: 0.33 °
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:

ALS

/ Beamline: 5.0.3 / Wavelength: 1 Å

Detector

Type: ADSC QUANTUM 315r / Detector: CCD / Date: Apr 21, 2022

Radiation

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

Radiation wavelength

Wavelength: 1 Å / Relative weight: 1

Reflection

Resolution: 2.83→47.43 Å / Num. obs: 115889 / % possible obs: 98.5 % / Redundancy: 3.5 % / B_iso Wilson estimate: 76.73 Å² / CC_1/2: 0.997 / R_merge(I) obs: 0.118 / R_pim(I) all: 0.073 / R_rim(I) all: 0.139 / Net I/σ(I): 6.1 / Num. measured all: 408951

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
2.83-2.88	3.3	1.789	19213	5736	0.297	1.169	2.149	0.5	98
15.5-47.43	3.6	0.027	2457	679	0.998	0.017	0.032	28.1	95.8

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Processing

Software

Name	Version	Classification
XDS		data reduction
Aimless	0.7.4	data scaling
PHENIX	1.19	refinement
PDB_EXTRACT	3.27	data extraction
PHENIX	1.19	phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 8DH0

8dh0

Resolution: 2.85→47.43 Å / SU ML: 0.51 / Cross valid method: THROUGHOUT / σ(F): 1.96 / Phase error: 33.24 / Stereochemistry target values: ML

	R_factor	Num. reflection	% reflection
R_free	0.2672	1951	1.72 %
R_work	0.2335	111411	-
obs	0.2341	113362	98.44 %

Solvent computation

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

Displacement parameters

B_iso max: 398.81 Å² / B_iso mean: 108.6907 Å² / B_iso min: 28.22 Å²

Refinement step

Cycle: final / Resolution: 2.85→47.43 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	25216	2652	126	0	27994
B_iso mean	-	-	107.66	-	-
Num. residues	-	-	-	-	3367

LS refinement shell

Refine-ID: X-RAY DIFFRACTION / R_factor R_free error: 0 / Total num. of bins used: 14

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Num. reflection all	% reflection obs (%)
2.85-2.92	0.4335	120	0.4054	7876	7996	97
2.92-3	0.3846	153	0.3511	7900	8053	98
3-3.09	0.3493	146	0.3198	7931	8077	98
3.09-3.19	0.3599	137	0.3047	7951	8088	98
3.19-3.3	0.329	131	0.2955	7953	8084	98
3.3-3.43	0.335	151	0.2957	7940	8091	98
3.43-3.59	0.3218	137	0.2535	7985	8122	98
3.59-3.78	0.3024	126	0.2276	7961	8087	99
3.78-4.02	0.3043	167	0.2266	7886	8053	98
4.02-4.33	0.2748	124	0.2112	8045	8169	99
4.33-4.76	0.2277	136	0.1951	7967	8103	99
4.76-5.45	0.26	146	0.207	8009	8155	99
5.45-6.86	0.2599	138	0.2333	8001	8139	99
6.86-47.43	0.1743	139	0.1915	8006	8145	99

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	1.8888	0.3783	0.2088	0.3657	-0.1233	1.0336	0.0801	0.9849	0.5108	-0.1606	-0.1132	-0.1497	-0.0418	0.3008	0.0434	0.5038	0.1129	0.0836	0.8869	0.3273	0.7339	32.9763	53.8532	-56.361
2	1.5612	-0.2398	-0.0264	1.3821	0.0666	0.9822	0.0135	-0.1001	0.1759	0.2127	-0.0294	-0.0242	-0.0254	-0.0414	0.045	0.2675	-0.0052	0.015	0.4068	-0.0057	0.3766	9.4064	41.769	-28.4599
3	1.9684	-0.1669	-0.0551	1.3143	-0.1969	1.7097	0.1879	0.6136	-0.1365	-0.1476	-0.1563	-0.217	0.2516	0.3868	0.0335	0.302	0.0487	0.0569	0.416	-0.0045	0.3744	25.6294	23.8563	-39.5297
4	2.5581	2.178	0.1278	1.9334	0.399	3.228	0.1768	0.4331	-0.0891	-0.6777	-0.1278	-1.1997	0.3978	1.1667	-0.2365	0.4178	0.1293	0.1168	0.9061	0.1194	0.8285	34.8419	34.7931	-46.251
5	4.0523	3.9898	-4.3005	4.2555	-4.4659	4.7318	-0.0942	2.0097	0.2421	-1.3583	0.0703	-0.5426	0.6032	0.3687	0.0078	0.542	0.0171	0.0999	0.8365	0.0555	0.5771	27.2039	44.0904	-41.3088
6	3.4305	1.0818	-0.966	0.5319	-0.1567	0.4021	-0.1648	0.7002	-0.1748	-0.9037	-0.3347	-0.4111	0.5817	-0.6886	0.5605	2.8714	0.9491	-0.8609	2.1008	-0.4972	2.2902	43.0702	21.6452	-47.7102
7	1.9401	0.1848	0.3523	2.3487	0.2312	0.9407	0.052	-0.7532	-0.6785	0.4053	0.0047	-0.4287	0.3451	-0.1499	0.0448	0.6273	-0.0131	0.0601	0.9171	0.1507	0.7297	32.2565	58.2871	8.8694
8	2.35	-0.0218	0.1984	1.2949	0.047	0.9604	-0.0462	-0.6273	0.0359	-0.0176	-0.0887	-0.0638	-0.0838	-0.047	0.1322	0.469	0.1149	0.0095	0.5034	-0.1428	0.4306	37.4037	82.9867	-6.3315
9	1.1356	-0.1272	1.5013	0.046	0.0435	3.4222	0.5655	-1.3388	1.0961	0.5204	0.534	-3.9482	-0.2214	3.844	-0.8401	1.2581	0.2013	0.0792	3.1874	-0.293	2.9372	72.9508	75.6844	2.8103
10	1.6595	-0.1868	-3.5134	7.0998	2.2809	7.9431	-0.3818	-1.2902	-0.6331	0.3091	-0.3167	-1.0884	0.0518	2.314	0.497	0.8658	0.2544	-0.0967	1.2805	0.1593	0.8251	50.2623	72.7923	-0.6416
11	1.9646	3.7364	0.7206	8.9768	-0.0545	5.1742	-0.0447	-0.3541	-1.7375	-0.8787	-0.0367	-1.4136	1.2741	1.5733	0.3267	0.8147	0.2559	0.1624	0.9759	0.0992	0.9385	32.8747	61.3222	3.515
12	2.7875	-1.8932	-4.5357	2.1304	3.2484	7.7403	-0.1742	-0.8937	-1.1629	-0.5476	-0.3871	-0.7759	1.0551	1.6056	0.7423	0.7883	0.0658	0.0759	1.0618	0.0698	0.9206	36.6685	64.7524	-1.9154
13	2.2423	-0.295	-0.9707	0.6407	0.5976	0.7852	-0.9123	-0.367	-0.1525	0.0634	-0.4532	-0.8085	-0.1797	-0.6455	1.2626	2.182	0.1086	0.21	1.8907	-0.3192	2.4385	64.7767	75.8163	2.578
14	2.1858	0.0037	-0.2046	0.7922	-0.1097	1.6817	-0.0267	-0.0386	0.8712	0.0577	0.1543	0.1627	-0.4445	-0.4694	-0.1254	0.687	-0.0308	0.0716	0.5378	0.0172	0.8992	-2.1485	45.9218	-95.4235
15	2.7201	-0.0009	0.124	1.5635	-0.3631	1.7985	0.3582	-0.3825	-0.4404	-0.2496	-0.0441	0.426	0.5667	-0.282	-0.2689	0.9122	-0.2127	-0.2617	0.6692	0.0677	0.5755	8.4027	10.3958	-91.21
16	1.0913	0.9221	0.8797	4.245	-0.3511	4.8171	1.0238	-0.551	1.3101	1.1088	-0.0744	1.9466	-0.5828	-1.5488	-0.8298	0.9556	-0.1838	0.1552	1.3278	0.2635	1.2893	-6.5755	23.2337	-88.4125
17	6.4949	-0.1827	-0.9691	1.0673	-2.4764	6.034	0.6018	0.1149	0.456	0.4244	0.1248	0.8144	1.0062	-0.9601	-0.7726	0.8495	-0.114	-0.1494	0.6654	0.0133	0.9931	3.1486	22.8695	-97.8879
18	0.0153	-0.004	0.0167	0.0016	0.0022	0.0089	-0.0329	0.9607	-0.4762	0.6628	-0.7766	0.6718	-0.306	-0.0686	0.7075	2.5959	-0.4365	-0.8763	3.5646	0.1588	3.5124	-17.9675	17.2941	-77.9789
19	4.7957	2.2107	-2.3424	1.3049	-1.3403	2.1906	-0.069	0.7558	-0.1271	-0.1283	0.4351	0.0449	0.9344	-0.5119	-0.4636	1.3844	0.1269	-0.3707	1.1505	-0.1316	0.8955	7.5068	-14.0623	54.6877
20	2.1468	0.057	1.1995	0.9859	0.2695	2.4761	0.1531	0.5117	0.1134	-0.0779	-0.2552	0.0913	-0.1557	0.2568	0.1116	0.9407	0.0908	-0.0483	0.9721	-0.087	0.4582	19.7847	19.3256	50.1212
21	1.3142	-1.8218	-2.1812	2.5496	3.0459	3.6398	1.2707	1.9216	1.2862	-2.8311	-0.3959	1.1688	-1.0891	-2.2977	-0.7422	3.0445	1.2191	-0.7612	3.1737	-0.7511	2.8513	-19.6354	24.0692	42.7763
22	9.9142	-4.485	1.7064	5.023	0.7777	1.0942	-0.508	0.3256	-0.7581	0.6809	-0.1421	1.1849	0.373	0.0478	0.6014	1.2102	0.0974	0.1451	1.3313	0.0331	0.5929	6.6233	10.0309	54.0153
23	5.5013	3.3812	5.3271	3.0995	3.4241	5.1966	1.0044	-0.952	0.1878	0.7622	-0.4463	1.0607	-0.2799	-0.5787	-0.4264	1.4142	0.0853	-0.0235	1.3929	-0.2629	0.7802	11.0857	9.0986	58.7422
24	3.6114	0.0866	-2.0844	0.7567	-0.7405	1.8281	-0.1449	0.5382	0.1081	0.0599	0.7237	0.4447	-1.1414	-1.6441	-0.6023	2.4296	0.6361	0.0617	2.6304	-0.3724	2.5259	-12.0755	22.9023	43.9059

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 1 through 331 )	B	1 - 331
2	X-RAY DIFFRACTION	2	chain 'B' and (resid 332 through 569 )	B	332 - 569
3	X-RAY DIFFRACTION	3	chain 'B' and (resid 570 through 883 )	B	570 - 883
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 3 through 18 )	A	3 - 18
5	X-RAY DIFFRACTION	5	chain 'C' and (resid 0 through 8 )	C	0 - 8
6	X-RAY DIFFRACTION	6	chain 'D' and (resid 2 through 8 )	D	2 - 8
7	X-RAY DIFFRACTION	7	chain 'E' and (resid 3 through 437 )	E	3 - 437
8	X-RAY DIFFRACTION	8	chain 'E' and (resid 438 through 883 )	E	438 - 883
9	X-RAY DIFFRACTION	9	chain 'F' and (resid 2 through 6 )	F	2 - 6
10	X-RAY DIFFRACTION	10	chain 'F' and (resid 7 through 12 )	F	7 - 12
11	X-RAY DIFFRACTION	11	chain 'F' and (resid 13 through 18 )	F	13 - 18
12	X-RAY DIFFRACTION	12	chain 'G' and (resid 1 through 8 )	G	1 - 8
13	X-RAY DIFFRACTION	13	chain 'H' and (resid 2 through 9 )	H	2 - 9
14	X-RAY DIFFRACTION	14	chain 'I' and (resid 2 through 263 )	I	2 - 263
15	X-RAY DIFFRACTION	15	chain 'I' and (resid 264 through 883 )	I	264 - 883
16	X-RAY DIFFRACTION	16	chain 'J' and (resid 3 through 18 )	J	3 - 18
17	X-RAY DIFFRACTION	17	chain 'K' and (resid 1 through 8 )	K	1 - 8
18	X-RAY DIFFRACTION	18	chain 'L' and (resid 2 through 8 )	L	2 - 8
19	X-RAY DIFFRACTION	19	chain 'M' and (resid 1 through 284 )	M	1 - 284
20	X-RAY DIFFRACTION	20	chain 'M' and (resid 285 through 882 )	M	285 - 882
21	X-RAY DIFFRACTION	21	chain 'N' and (resid 2 through 6 )	N	2 - 6
22	X-RAY DIFFRACTION	22	chain 'N' and (resid 7 through 18 )	N	7 - 18
23	X-RAY DIFFRACTION	23	chain 'O' and (resid 1 through 8 )	O	1 - 8
24	X-RAY DIFFRACTION	24	chain 'P' and (resid 2 through 9 )	P	2 - 9

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