PDB-8bgj: Crystal structure of Reverse Transcriptase domain from Caloramato...

ID or keywords:

Entry

Database: PDB / ID: 8bgj

Title

Crystal structure of Reverse Transcriptase domain from Caloramator australicus CART-CAPP

Components

RNA-directed DNA polymerase

Keywords

TRANSFERASE / Reverse transcriptase / CRISPR associated reverse transcriptase / CART

Function / homology

Function and homology information

RNA-directed DNA polymerase / RNA-directed DNA polymerase activity / defense response to virus / RNA binding
Similarity search - Function

Biological species

Caloramator australicus RC3 (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.63 Å

Authors

Li, A.W.H. / Doherty, A.J.

Funding support

United Kingdom, 3items

Organization	Grant number	Country
Biotechnology and Biological Sciences Research Council (BBSRC)	BB/S008691/1	United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)	BB/W015226/1	United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)	BB/P007031/1	United Kingdom

Citation

Journal: Nucleic Acids Res. / Year: 2023
Title: Reverse transcriptases prime DNA synthesis.
Authors: Zabrady, M. / Zabrady, K. / Li, A.W.H. / Doherty, A.J.

History

Deposition	Oct 27, 2022	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Jun 7, 2023	Provider: repository / Type: Initial release
Revision 1.1	Jun 21, 2023	Group: Database references / Category: citation / citation_author Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID
Revision 1.2	Aug 23, 2023	Group: Data collection / Database references Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / citation / citation_author Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.3	Feb 7, 2024	Group: Refinement description / Category: pdbx_initial_refinement_model

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	8bgj.cif.gz	329.4 KB	Display	PDBx/mmCIF format
PDB format	pdb8bgj.ent.gz	222.8 KB	Display	PDB format
PDBx/mmJSON format	8bgj.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	8bgj_validation.pdf.gz	484 KB	Display	wwPDB validaton report
Full document	8bgj_full_validation.pdf.gz	499.9 KB	Display
Data in XML	8bgj_validation.xml.gz	28.8 KB	Display
Data in CIF	8bgj_validation.cif.gz	40.4 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/bg/8bgj ftp://data.pdbj.org/pub/pdb/validation_reports/bg/8bgj	HTTPS FTP

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Related structure data

Related structure data	8baoC 6ar1S 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
Related items in Molecule of the Month	#227 - Nov 2018 Telomerase similarity (1) #228 - Dec 2018 Directed Evolution of Enzymes similarity (1)

Deposited unit

A: RNA-directed DNA polymerase

B: RNA-directed DNA polymerase

C: RNA-directed DNA polymerase

hetero molecules

81.4 kDa, 3 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: RNA-directed DNA polymerase

hetero molecules

defined by author
Evidence: gel filtration
27.3 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

B: RNA-directed DNA polymerase

hetero molecules

defined by author
Evidence: gel filtration
27.2 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

3

C: RNA-directed DNA polymerase

hetero molecules

defined by author
Evidence: gel filtration
26.9 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

Unit cell

Length a, b, c (Å)	96.920, 96.920, 65.400
Angle α, β, γ (deg.)	90.000, 90.000, 120.000
Int Tables number	145
Space group name H-M	P3₂
Space group name Hall	P3₂
Symmetry operation	#1: x,y,z #2: -y,x-y,z+2/3 #3: -x+y,-x,z+1/3

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Protein , 1 types, 3 molecules ABC

#1: Protein	RNA-directed DNA polymerase Mass: 26569.090 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Caloramator australicus RC3 (bacteria) / Gene: CAAU_1036 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: I7LG99, RNA-directed DNA polymerase

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Non-polymers , 5 types, 344 molecules

#2: Chemical	ChemComp-MG / MAGNESIUM ION Mass: 24.305 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Mg
#3: Chemical	ChemComp-1PE / PENTAETHYLENE GLYCOL / PEG400 Mass: 238.278 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C₁₀H₂₂O₆ / Comment: precipitant*YM
#4: Chemical	ChemComp-NA / SODIUM ION Mass: 22.990 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Na
#5: Chemical	ChemComp-TLA / L(+)-TARTARIC ACID Mass: 150.087 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₄H₆O₆
#6: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 331 / Source method: isolated from a natural source / Formula: H₂O

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Details

Has ligand of interest	N

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Experiment

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

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Sample preparation

Crystal	Density Matthews: 2.22 Å³/Da / Density % sol: 44.72 %
Crystal grow	Temperature: 287.15 K / Method: vapor diffusion, sitting drop / Details: PEG 3350, Sodium tartrate dibasic dihydrate

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: Diamond / Beamline: I24 / Wavelength: 0.9686 Å
Detector	Type: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Feb 20, 2020
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 0.9686 Å / Relative weight: 1
Reflection	Resolution: 1.63→83.94 Å / Num. obs: 85450 / % possible obs: 99.6 % / Redundancy: 3.1 % / B_iso Wilson estimate: 25.71 Å² / CC_1/2: 0.992 / R_merge(I) obs: 0.076 / R_pim(I) all: 0.051 / R_rim(I) all: 0.092 / Net I/σ(I): 9
Reflection shell	Resolution: 1.63→1.66 Å / Redundancy: 3.1 % / R_merge(I) obs: 1.184 / Mean I/σ(I) obs: 0.5 / Num. unique obs: 4204 / CC_1/2: 0.318 / R_pim(I) all: 0.798 / R_rim(I) all: 1.432 / % possible all: 99.5

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Processing

Software

Name	Version	Classification
xia2		data reduction
PHENIX	1.20.1_4487	refinement
DIALS		data scaling
PHASER		phasing
Coot		model building

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 6ar1

6ar1

Resolution: 1.63→48.46 Å / SU ML: 0.288 / Cross valid method: FREE R-VALUE / σ(F): 1.96 / Phase error: 23.3426
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2

	R_factor	Num. reflection	% reflection
R_free	0.2188	4182	4.9 %
R_work	0.183	81105	-
obs	0.1846	85287	99.45 %

Solvent computation

Shrinkage radii: 0.9 Å / VDW probe radii: 1.1 Å / Solvent model: FLAT BULK SOLVENT MODEL

Displacement parameters

B_iso mean: 41.23 Å²

Refinement step

Cycle: LAST / Resolution: 1.63→48.46 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4954	0	112	331	5397

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.0073	5277
X-RAY DIFFRACTION	f_angle_d	0.9495	7070
X-RAY DIFFRACTION	f_chiral_restr	0.0633	760
X-RAY DIFFRACTION	f_plane_restr	0.0082	891
X-RAY DIFFRACTION	f_dihedral_angle_d	15.6752	738

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 (%)
1.63-1.65	0.4107	138	0.4111	2637	X-RAY DIFFRACTION	97.54
1.65-1.67	0.4214	155	0.4108	2671	X-RAY DIFFRACTION	98.12
1.67-1.69	0.376	150	0.3952	2654	X-RAY DIFFRACTION	97.87
1.69-1.71	0.3856	152	0.3763	2649	X-RAY DIFFRACTION	98.56
1.71-1.73	0.3632	146	0.3433	2705	X-RAY DIFFRACTION	98.79
1.73-1.76	0.3359	105	0.32	2700	X-RAY DIFFRACTION	99.64
1.76-1.78	0.3134	174	0.3096	2662	X-RAY DIFFRACTION	99.02
1.78-1.81	0.3236	147	0.2726	2688	X-RAY DIFFRACTION	99.33
1.81-1.84	0.2642	134	0.2588	2685	X-RAY DIFFRACTION	99.33
1.84-1.87	0.272	187	0.229	2710	X-RAY DIFFRACTION	99.42
1.87-1.9	0.2268	156	0.2174	2686	X-RAY DIFFRACTION	99.72
1.9-1.93	0.2606	138	0.2193	2709	X-RAY DIFFRACTION	99.72
1.93-1.97	0.2476	150	0.2135	2681	X-RAY DIFFRACTION	99.89
1.97-2.01	0.2355	165	0.2073	2682	X-RAY DIFFRACTION	99.62
2.01-2.05	0.2631	124	0.2086	2751	X-RAY DIFFRACTION	99.48
2.05-2.1	0.2303	154	0.2181	2690	X-RAY DIFFRACTION	99.89
2.1-2.15	0.1884	134	0.1666	2708	X-RAY DIFFRACTION	99.44
2.15-2.21	0.1912	179	0.1631	2664	X-RAY DIFFRACTION	99.51
2.21-2.28	0.1822	136	0.1614	2706	X-RAY DIFFRACTION	99.89
2.28-2.35	0.2184	124	0.173	2721	X-RAY DIFFRACTION	99.89
2.35-2.43	0.2251	144	0.1727	2764	X-RAY DIFFRACTION	99.97
2.43-2.53	0.218	122	0.1736	2696	X-RAY DIFFRACTION	99.75
2.53-2.65	0.2159	148	0.1748	2724	X-RAY DIFFRACTION	99.97
2.65-2.79	0.2147	100	0.1863	2756	X-RAY DIFFRACTION	99.96
2.79-2.96	0.1929	115	0.1757	2750	X-RAY DIFFRACTION	100
2.96-3.19	0.2072	110	0.1658	2750	X-RAY DIFFRACTION	99.93
3.19-3.51	0.1994	132	0.1644	2695	X-RAY DIFFRACTION	99.86
3.51-4.02	0.1759	140	0.143	2740	X-RAY DIFFRACTION	100
4.02-5.06	0.1668	119	0.1381	2721	X-RAY DIFFRACTION	99.51
5.06-48.46	0.2354	104	0.1725	2750	X-RAY DIFFRACTION	99.79

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	5.56285786808	1.85217167445	-0.712310044177	5.43586666246	1.08319897497	2.94874897881	-0.595860822854	-1.72413030496	-0.152369633681	1.33388030171	0.192759299034	0.316550672334	1.85706933118	-1.29486022779	0.271710529929	1.28342780795	-0.157765733895	0.158469717535	1.00271022196	0.10309246564	1.03497877614	-2.35967888284	35.8098177612	8.78834797146
2	1.3373025454	1.27839978941	-0.044546012204	5.36549010746	-1.48807965454	2.45732847417	-0.160018975283	0.127550065401	0.223061373309	-0.307084568579	0.18381089233	0.194754251316	-0.316738121468	-0.142491721722	-0.0437868053054	0.247996271092	0.0268961605907	-0.0147604940091	0.222879436924	-0.00759996384198	0.174136761674	-0.0248053800237	17.8768615007	-3.54255240776
3	0.721234515902	0.224801897572	-0.153079890115	6.05321378568	-1.7426624721	0.808190848928	-0.0281610377769	0.0331673476483	0.14575435807	0.120525898408	-0.0210499562443	-0.439936474543	-0.127196908761	0.0660036924688	0.0535705591836	0.197821355555	0.00760993373483	-0.0211817570344	0.180660647276	-0.0211405001297	0.226096363664	10.0618971772	19.6182910878	2.18324057131
4	3.05236894734	2.08786901515	-0.617129084727	4.53969611365	-2.59917340278	2.21355184542	-0.0257247379639	-0.202072345456	0.0378243847173	0.530339633866	-0.0997943603939	-0.302249622197	-0.413527120577	0.0370447885714	0.108045304385	0.31611270384	0.0416359849332	-0.0402339082214	0.219578497934	-0.0283452413302	0.199322572602	5.40666175903	8.706760363	13.7169417095
5	5.26586445933	0.617295750264	-0.876719888356	6.61196261271	-2.43128586067	3.99422853563	-0.0939838025807	-0.304590997272	-0.872383641901	0.238008299112	-0.0618010005906	-0.443320003323	0.301729490348	0.522942581011	0.104397789528	0.307113328964	-0.0136620548979	-0.0340648815602	0.414771397059	-0.0740783217594	0.404843015957	22.6180558289	6.77100057027	-28.0863826968
6	2.51268230988	0.794826001025	0.498267451117	1.02309609511	0.289392400174	0.778519133567	-0.00155821566882	-0.0088680721701	0.0310266539334	0.020544052012	-0.0429282557276	0.166020348114	0.038965301957	-0.130002177442	0.0639745115732	0.189515296996	-0.00665959239133	0.0145766345271	0.185700669376	-0.0185377133201	0.178154591246	30.1039250584	22.3405703452	-23.1835076195
7	6.32782669933	0.453398737833	2.3713895313	2.01579533211	1.22298750031	2.91352713593	0.13362429455	-0.363822046165	0.235383159243	0.189741227135	-0.21816847718	0.156263853022	0.19972846366	-0.358327206993	0.11017090393	0.258474994186	-0.0447717641851	0.0442778096777	0.257630503324	-0.0225958735726	0.190294080113	38.0983140246	26.509556859	-11.132764164
8	3.89837702272	-0.11788819565	-1.61826368717	6.0429542607	4.20073160494	3.51223255023	-0.184596997725	0.581582172057	0.558675879182	-1.18119539993	0.747441295988	0.783348037772	-0.0527962988082	-1.75023087116	-0.628309277512	1.08353514832	-0.176216210347	-0.170110136276	1.48537631028	0.291769207634	1.59032933371	29.0729014417	36.1703855936	0.59830114477
9	4.20896249045	0.317318520502	-0.589726437379	2.05227234761	-0.0933759590098	3.1807676846	-0.0430239291919	-0.635116788376	-0.315255945759	0.157493630629	-0.0278080559851	-0.313095085872	0.412410306725	0.559544495299	-0.0912794114573	0.25816763928	0.0774055275998	0.0164654509373	0.280022238727	0.047355542971	0.180483448081	14.6660820663	47.7995662472	14.2887779988
10	4.47314049367	-1.66302196195	-0.677840754539	1.82853251574	0.382356151267	0.914598302646	-0.145974402113	-0.46334339663	0.234234914055	0.117645774378	0.234484789516	-0.315017752565	0.0764880898985	0.372949946534	-0.0544602149762	0.206333121637	0.0383087081867	0.00687551258851	0.306293624124	-0.0300546948798	0.215691617943	22.233921985	54.3145903722	8.08129420887
11	4.77625175704	-0.36137036784	2.34010753818	1.61078512586	-0.712279540543	2.3927979346	0.0712328436517	0.53368563865	0.242654018037	-0.294483140461	-0.118593791342	-0.188631676627	0.160271621464	0.356859673742	-0.0122674256121	0.220554230945	0.0404945274537	0.054747753642	0.282421397188	0.0186250264943	0.214403594955	10.0375683486	56.1871852514	-2.9712091625

Refinement TLS group

Refine-ID: X-RAY DIFFRACTION

ID	Refine TLS-ID	Selection details	Auth asym-ID	Label asym-ID	Auth seq-ID	Label seq-ID
1	1	chain 'A' and (resid 1 through 20 )	A	A	1 - 20	1 - 20
2	2	chain 'A' and (resid 21 through 65 )	A	A	21 - 65	21 - 65
3	3	chain 'A' and (resid 66 through 164 )	A	A	66 - 164	66 - 164
4	4	chain 'A' and (resid 165 through 203 )	A	A	165 - 203	165 - 203
5	5	chain 'B' and (resid 12 through 36 )	B	G	12 - 36	1 - 25
6	6	chain 'B' and (resid 37 through 164 )	B	G	37 - 164	26 - 153
7	7	chain 'B' and (resid 165 through 203 )	B	G	165 - 203	154 - 192
8	8	chain 'C' and (resid 1 through 23 )	C	M	1 - 23	1 - 23
9	9	chain 'C' and (resid 24 through 65 )	C	M	24 - 65	24 - 65
10	10	chain 'C' and (resid 66 through 164 )	C	M	66 - 164	66 - 164
11	11	chain 'C' and (resid 165 through 203 )	C	M	165 - 203	165 - 203

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