PDB-8dh4: T7 RNA polymerase elongation complex with unnatural base dPa-DsTP pair

ID or keywords:

Entry

Database: PDB / ID: 8dh4

Title

T7 RNA polymerase elongation complex with unnatural base dPa-DsTP pair

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 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / 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: 2.8 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	8dh4.cif.gz	1.4 MB	Display	PDBx/mmCIF format
PDB format	pdb8dh4.ent.gz	1.2 MB	Display	PDB format
PDBx/mmJSON format	8dh4.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	8dh4_validation.pdf.gz	1.2 MB	Display	wwPDB validaton report
Full document	8dh4_full_validation.pdf.gz	1.2 MB	Display
Data in XML	8dh4_validation.xml.gz	114.6 KB	Display
Data in CIF	8dh4_validation.cif.gz	155.6 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/dh/8dh4 ftp://data.pdbj.org/pub/pdb/validation_reports/dh/8dh4	HTTPS FTP

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Related structure data

Related structure data	8dh0SC 8dh1C 8dh2C 8dh3C 8dh5C 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

445 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

defined by author
111 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

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

Unit cell

Length a, b, c (Å)	78.779, 86.230, 201.463
Angle α, β, γ (deg.)	89.850, 85.390, 69.590
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, 4 molecules

#5: Chemical	ChemComp-S96 / (7P)-3-{5-O-[(R)-hydroxy{[(S)-hydroxy(phosphonooxy)phosphoryl]oxy}phosphoryl]-beta-D-ribofuranosyl}-7-(thiophen-2-yl)-3H-imidazo[4,5-b]pyridine Mass: 573.302 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₁₅H₁₈N₃O₁₃P₃S / Feature type: SUBJECT OF INVESTIGATION
#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: 2.88 Å³/Da / Density % sol: 57.26 % / Mosaicity: 0.19 °
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: 8.2.1 / Wavelength: 1 Å

Detector

Type: ADSC QUANTUM 315r / Detector: CCD / Date: Dec 9, 2021

Radiation

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

Radiation wavelength

Wavelength: 1 Å / Relative weight: 1

Reflection

Resolution: 2.75→48.84 Å / Num. obs: 125883 / % possible obs: 97.8 % / Redundancy: 3.8 % / B_iso Wilson estimate: 82.06 Å² / CC_1/2: 0.995 / R_merge(I) obs: 0.099 / R_pim(I) all: 0.059 / R_rim(I) all: 0.115 / Net I/σ(I): 8.2 / Num. measured all: 481832 / Scaling rejects: 17

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
2.75-2.8	3.9	2.239	24587	6284	0.253	1.31	2.595	0.6	97.8
15.06-48.84	3.7	0.037	2503	668	0.973	0.024	0.044	29	85.4

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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: 2.8→43.2 Å / SU ML: 0.5 / Cross valid method: THROUGHOUT / σ(F): 1.96 / Phase error: 33.9 / Stereochemistry target values: ML

	R_factor	Num. reflection	% reflection
R_free	0.2745	1905	1.6 %
R_work	0.2328	117218	-
obs	0.2334	119123	97.72 %

Solvent computation

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

Displacement parameters

B_iso max: 328.61 Å² / B_iso mean: 116.8149 Å² / B_iso min: 32.2 Å²

Refinement step

Cycle: final / Resolution: 2.8→43.2 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	25963	2411	72	0	28446
B_iso mean	-	-	111.28	-	-
Num. residues	-	-	-	-	3432

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 (%)
2.8-2.87	0.4803	152	0.4513	8332	8484	98
2.87-2.95	0.4115	122	0.3855	8402	8524	98
2.95-3.03	0.3657	139	0.3328	8407	8546	98
3.03-3.13	0.3564	150	0.3039	8357	8507	98
3.13-3.24	0.3422	135	0.3035	8399	8534	98
3.24-3.37	0.3447	121	0.3065	8404	8525	98
3.37-3.53	0.3376	144	0.2571	8400	8544	98
3.53-3.71	0.3093	130	0.2332	8442	8572	98
3.71-3.95	0.3012	145	0.232	8459	8604	98
3.95-4.25	0.2445	129	0.2197	8371	8500	98
4.25-4.68	0.251	136	0.1959	8408	8544	98
4.68-5.35	0.2362	134	0.2022	8376	8510	98
5.35-6.74	0.2441	137	0.2292	8326	8463	97
6.74-43.2	0.2208	131	0.1891	8135	8266	95

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	1.8968	0.3129	-0.0342	0.0411	-0.1199	1.0319	0.1635	1.1346	0.5345	-0.0738	-0.1477	-0.1437	-0.0906	0.2061	-0.0039	0.609	0.0731	0.073	1.0732	0.4368	0.9199	33.0579	54.1132	-56.4302
2	1.9768	-0.2717	0.354	1.0992	-0.311	1.1926	0.1482	0.3071	-0.0761	0.0076	-0.1379	-0.1649	0.2211	0.0827	-0.0233	0.3296	-0.0229	0.0201	0.4702	0.0372	0.3472	18.7129	31.5067	-34.7474
3	2.4835	1.9903	0.6359	1.6299	0.581	4.2288	0.235	0.3981	-0.0333	-0.555	0.0165	-1.7329	0.4348	1.3645	-0.7327	0.4758	-0.0002	0.1185	1.2205	0.1877	1.1065	33.9311	36.3091	-45.2111
4	4.2468	5.611	-2.7199	8.4292	-2.4199	3.1795	0.4113	1.6599	0.528	-0.6849	0.1175	-0.5022	0.5496	0.4442	-0.2916	0.5036	-0.0204	0.0442	1.0292	0.2114	0.5329	27.271	44.2334	-41.3173
5	5.1068	0.9612	1.9872	7.5579	-0.0941	1.5526	0.5504	0.7963	-0.0708	-1.7619	-0.0514	-1.394	0.5634	-0.1435	-0.499	3.289	0.1965	-0.2106	2.709	-0.0394	2.689	42.1688	20.5084	-45.9494
6	2.2626	0.288	0.6896	2.0127	0.0555	1.4775	0.0779	-0.7751	-0.8079	0.2095	-0.0469	-0.6517	0.4376	0.0736	-0.0804	0.6787	0.0371	0.1017	1.0251	0.2297	0.9915	35.9707	55.8144	8.7629
7	2.5931	0.2308	0.0241	1.3951	-0.0423	0.8616	-0.0372	-0.6586	0.0848	-0.0112	-0.1337	-0.0486	-0.1199	-0.097	0.1577	0.4595	0.063	-0.0152	0.5867	-0.1081	0.3948	36.1599	82.641	-5.6354
8	0.1341	-0.5334	-0.6265	2.1538	2.532	2.9981	0.4423	-0.5177	-0.0662	0.1883	0.1659	-1.9406	-0.5486	1.1184	-0.7271	2.5783	0.1873	-0.1317	3.3809	-0.6067	3.0403	72.743	74.6646	1.7737
9	4.4786	-1.827	-2.3002	9.8035	1.0538	2.0035	-0.0296	-0.6564	-0.5053	0.2885	-0.2257	-1.6098	0.8025	2.482	0.3394	1.0026	0.1783	-0.2018	1.6615	0.0619	0.9397	50.1284	73.0229	-0.576
10	2.3723	4.2393	-0.2023	7.8351	1.1062	8.018	-0.3291	-1.4734	-1.9377	-0.2396	-0.5695	-1.4754	1.5232	0.9209	0.6053	1.0509	0.2089	0.2128	1.2513	0.3142	1.2346	32.7432	61.5536	3.6187
11	7.2786	-1.2994	-4.3296	2.6047	0.9166	2.7136	-0.5364	-1.2618	-0.4222	-0.521	0.2524	-1.5047	1.1828	1.4316	0.3387	1.0451	0.1386	0.0094	1.2299	0.0089	0.9803	36.6063	64.8622	-1.8219
12	0.6101	0.4202	-0.4278	1.8171	1.9279	3.5856	-0.6732	0.0531	0.4032	0	-0.0023	-0.5428	-0.9663	0.1639	0.7766	2.4525	-0.091	-0.5312	3.0725	-0.1336	2.9935	64.8222	76.1695	1.8565
13	3.983	-3.3728	3.5213	4.2302	-2.2168	3.7985	-0.4763	0.0605	1.6389	0.1223	-0.0016	-0.6857	-0.4809	0.6472	0.5893	0.6709	-0.2219	0.0167	0.6609	-0.0351	0.8747	27.1068	38.1795	-95.9683
14	3.1493	0.1545	-0.2985	0.8278	-0.1282	1.9214	0.1513	0.1289	1.2804	0.1741	0.1257	0.4949	-0.5149	-0.622	-0.1472	0.7817	0.0903	0.1297	0.6616	0.0647	1.1789	-5.8996	45.3368	-94.3244
15	2.5724	-0.1358	0.0242	1.3283	-0.4857	2.6814	0.2491	0.0132	-0.3553	-0.3305	-0.0199	-0.0009	0.6155	0.181	-0.146	0.9746	0.0033	-0.1302	0.6788	-0.0492	0.443	19.6305	10.997	-101.6863
16	3.5808	0.2336	0.431	1.9287	-0.0306	2.5592	0.3306	-0.4854	-0.9592	-0.5898	-0.1094	1.0758	0.4785	-0.4991	-0.1289	1.1982	-0.3341	-0.4813	1.002	0.2258	1.1831	-9.4767	5.02	-88.5066
17	3.5776	-0.6161	-0.1634	1.805	0.1358	2.3444	0.11	-1.1841	-0.1511	-0.0858	0.0547	0.2127	0.4619	0.1721	-0.1752	0.8687	-0.2015	-0.0862	0.8573	0.0623	0.5696	9.7445	11.5678	-79.1548
18	1.7727	0.2399	2.7084	4.125	1.264	4.4387	1.0076	-1.0758	0.6524	0.8746	-0.1132	2.2052	-0.3832	-1.2173	-1.0204	1.0962	-0.0282	0.1552	1.0071	0.1996	1.2155	-2.8703	24.186	-91.0566
19	5.9345	-2.5367	0.6444	4.029	-3.0204	2.9311	0.5313	-0.4327	0.5612	0.0112	0.4389	0.6697	1.0354	-0.61	-0.7283	1.0345	-0.1003	-0.1847	0.5292	-0.1226	0.9633	3.3126	23.253	-98.1645
20	6.0966	-1.5977	3.5038	1.606	-3.3942	7.2778	-0.3519	1.0841	-0.8894	-0.7447	-1.15	1.6258	0.7835	-1.0835	1.2881	3.8824	0.5094	-0.7759	2.8241	-0.4723	3.087	-16.79	16.7243	-76.5146
21	4.0542	1.8085	-1.5499	1.4064	-0.9893	2.7663	0.2902	0.4944	-0.1651	0.0933	0.3098	0.5105	0.5777	-0.7586	-0.4383	1.1745	0.0357	-0.1908	1.1942	0.0864	1.0243	-0.0884	-12.0274	58.309
22	2.3116	-0.0422	1.0107	0.8315	0.1421	2.337	0.0268	0.5367	0.2776	-0.0304	-0.1483	0.103	-0.2923	0.2598	0.1324	1.1094	0.0462	-0.0642	1.0172	-0.0375	0.5519	19.898	19.6963	50.1302
23	7.0152	-1.3249	-1.8675	2.3102	1.2512	1.1821	0.1911	1.2261	-0.4394	0.1092	-0.4217	1.5216	-0.1996	-0.8538	0.2474	1.5284	0.2159	0.0679	1.7248	-0.0372	1.0869	1.5021	12.9926	52.5321
24	2.5259	-1.7369	-1.4509	4.9297	-2.6933	7.0631	0.3242	0.6733	-0.2528	-0.0171	0.2532	1.3023	-0.5564	0.3653	-0.4125	1.571	0.0804	-0.186	1.5151	-0.2314	0.8221	11.256	9.0132	58.6601
25	5.3418	0.5983	-2.6	3.7814	-3.2162	3.5728	-0.7284	0.2611	-0.358	0.4781	-0.7783	0.5077	1.051	-1.2468	1.3472	3.5375	0.3959	-0.2074	2.9672	-1.163	2.9392	-9.3052	25.3389	44.3836

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 883 )	B	332 - 883
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 4 through 18 )	A	4 - 18
4	X-RAY DIFFRACTION	4	chain 'C' and (resid 0 through 8 )	C	0 - 8
5	X-RAY DIFFRACTION	5	chain 'D' and (resid 2 through 7 )	D	2 - 7
6	X-RAY DIFFRACTION	6	chain 'E' and (resid 4 through 407 )	E	4 - 407
7	X-RAY DIFFRACTION	7	chain 'E' and (resid 408 through 883 )	E	408 - 883
8	X-RAY DIFFRACTION	8	chain 'F' and (resid 2 through 6 )	F	2 - 6
9	X-RAY DIFFRACTION	9	chain 'F' and (resid 7 through 12 )	F	7 - 12
10	X-RAY DIFFRACTION	10	chain 'F' and (resid 13 through 18 )	F	13 - 18
11	X-RAY DIFFRACTION	11	chain 'G' and (resid 1 through 8 )	G	1 - 8
12	X-RAY DIFFRACTION	12	chain 'H' and (resid 2 through 9 )	H	2 - 9
13	X-RAY DIFFRACTION	13	chain 'I' and (resid 3 through 61 )	I	3 - 61
14	X-RAY DIFFRACTION	14	chain 'I' and (resid 62 through 301 )	I	62 - 301
15	X-RAY DIFFRACTION	15	chain 'I' and (resid 302 through 549 )	I	302 - 549
16	X-RAY DIFFRACTION	16	chain 'I' and (resid 550 through 728 )	I	550 - 728
17	X-RAY DIFFRACTION	17	chain 'I' and (resid 729 through 883 )	I	729 - 883
18	X-RAY DIFFRACTION	18	chain 'J' and (resid 5 through 18 )	J	5 - 18
19	X-RAY DIFFRACTION	19	chain 'K' and (resid 1 through 8 )	K	1 - 8
20	X-RAY DIFFRACTION	20	chain 'L' and (resid 4 through 6 )	L	4 - 6
21	X-RAY DIFFRACTION	21	chain 'M' and (resid 3 through 283 )	M	3 - 283
22	X-RAY DIFFRACTION	22	chain 'M' and (resid 284 through 883 )	M	284 - 883
23	X-RAY DIFFRACTION	23	chain 'N' and (resid 4 through 18 )	N	4 - 18
24	X-RAY DIFFRACTION	24	chain 'O' and (resid 1 through 8 )	O	1 - 8
25	X-RAY DIFFRACTION	25	chain 'P' and (resid 2 through 7 )	P	2 - 7

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