PDB-5xuu: Crystal structure of Lachnospiraceae bacterium ND2006 Cpf1 in com...

ID or keywords:

Entry

Database: PDB / ID: 5xuu

Title

Crystal structure of Lachnospiraceae bacterium ND2006 Cpf1 in complex with crRNA and target DNA (TCCA PAM)

Components

DNA (29-MER)
DNA (5'-D(*CP*GP*TP*CP*CP*TP*CP*CP*A)-3')
LbCpf1
crRNA

Keywords

HYDROLASE/RNA/DNA / nuclease / HYDROLASE-RNA-DNA complex

Function / homology

Function and homology information

metal ion binding
Similarity search - Function

Biological species

Lachnospiraceae bacterium ND2006 (bacteria)
synthetic construct (others)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.5 Å

Authors

Yamano, T. / Nishimasu, H. / Ishitani, R. / Nureki, O.

Citation

Journal: Mol. Cell / Year: 2017
Title: Structural Basis for the Canonical and Non-canonical PAM Recognition by CRISPR-Cpf1.
Authors: Yamano, T. / Zetsche, B. / Ishitani, R. / Zhang, F. / Nishimasu, H. / Nureki, O.

History

Deposition	Jun 26, 2017	Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0	Aug 9, 2017	Provider: repository / Type: Initial release
Revision 1.1	Dec 6, 2017	Group: Database references / Category: citation Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2	Nov 22, 2023	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 / pdbx_struct_conn_angle / struct_conn Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_conn_angle.ptnr1_auth_asym_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.value / _struct_conn.pdbx_dist_value / _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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Related structure data

Related structure data	5xusSC 5xutC 5xuzC S: Starting model for refinement C: citing same article (ref.)
Similar structure data	5xus-d 6gtg-d 5xuz-2 5xuz-1 5kk5-d 5xut-d 6i1k-d 5mga-d 6i1l-1 6i1l-2 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
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PDB pages

PDBj /

wwPDB /

NCBI

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Deposited unit

A: LbCpf1

B: crRNA

C: DNA (29-MER)

D: DNA (5'-D(*CP*GP*TP*CP*CP*TP*CP*CP*A)-3')

hetero molecules

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

1

Idetical with deposited unit
defined by author&software
Evidence: gel filtration

Unit cell

Length a, b, c (Å)	101.954, 101.954, 372.544
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	92
Space group name H-M	P4₁2₁2

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DNA chain , 2 types, 2 molecules CD

#3: DNA chain	DNA (29-MER) / Target DNA strand Mass: 8904.736 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#4: DNA chain	DNA (5'-D(CPGPTPCPCPTPCPCPA)-3') / Non- target DNA strand Mass: 2651.750 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source*: (synth.) synthetic construct (others)

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Protein / RNA chain , 2 types, 2 molecules AB

#1: Protein	LbCpf1 Mass: 144170.625 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Lachnospiraceae bacterium ND2006 (bacteria) Production host: Escherichia coli (E. coli) / References: UniProt: A0A182DWE3*PLUS
#2: RNA chain	crRNA Mass: 12879.634 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)

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Non-polymers , 4 types, 23 molecules

#5: Chemical	ChemComp-MG / MAGNESIUM ION Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg
#6: Chemical	ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL Mass: 62.068 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₂H₆O₂
#7: Chemical	ChemComp-NA / SODIUM ION Mass: 22.990 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Na
#8: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 19 / 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: 2.87 Å³/Da / Density % sol: 57.16 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 5 / Details: PEG 3350, MIB buffer, pH 5.0

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Data collection

Diffraction	Mean temperature: 100 K
Diffraction source	Source: SYNCHROTRON / Site: SPring-8 / Beamline: BL41XU / Wavelength: 1 Å
Detector	Type: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Jul 26, 2016
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 1 Å / Relative weight: 1
Reflection	Resolution: 2.5→49.17 Å / Num. obs: 69125 / % possible obs: 99.8 % / Redundancy: 8.6 % / Net I/σ(I): 15.6
Reflection shell	Resolution: 2.5→2.53 Å

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Processing

Software

Name	Version	Classification
PHENIX	(1.11.1_2575)	refinement
DIALS		data reduction
Aimless		data scaling
MOLREP		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 5XUS

5xus

Resolution: 2.5→49.169 Å / SU ML: 0.4 / Cross valid method: FREE R-VALUE / σ(F): 1.33 / Phase error: 27.26

	R_factor	Num. reflection	% reflection
R_free	0.2443	3438	4.98 %
R_work	0.1927	-	-
obs	0.1952	69014	99.71 %

Solvent computation

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

Refinement step

Cycle: LAST / Resolution: 2.5→49.169 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	9737	1618	10	19	11384

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.007	11775
X-RAY DIFFRACTION	f_angle_d	0.924	16256
X-RAY DIFFRACTION	f_dihedral_angle_d	17.843	6812
X-RAY DIFFRACTION	f_chiral_restr	0.05	1802
X-RAY DIFFRACTION	f_plane_restr	0.005	1813

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 (%)
2.5-2.5343	0.3829	112	0.3395	2617	X-RAY DIFFRACTION	100
2.5343-2.5705	0.3647	131	0.3032	2552	X-RAY DIFFRACTION	100
2.5705-2.6089	0.3232	153	0.2919	2576	X-RAY DIFFRACTION	100
2.6089-2.6496	0.3501	136	0.2787	2568	X-RAY DIFFRACTION	100
2.6496-2.6931	0.3271	147	0.2696	2573	X-RAY DIFFRACTION	100
2.6931-2.7395	0.3619	140	0.2616	2587	X-RAY DIFFRACTION	100
2.7395-2.7893	0.3468	134	0.2535	2568	X-RAY DIFFRACTION	100
2.7893-2.8429	0.3155	129	0.2542	2608	X-RAY DIFFRACTION	100
2.8429-2.901	0.3396	128	0.2584	2548	X-RAY DIFFRACTION	99
2.901-2.964	0.3407	170	0.265	2554	X-RAY DIFFRACTION	100
2.964-3.033	0.3445	122	0.2803	2615	X-RAY DIFFRACTION	100
3.033-3.1088	0.2821	122	0.2625	2619	X-RAY DIFFRACTION	100
3.1088-3.1928	0.285	129	0.2379	2640	X-RAY DIFFRACTION	100
3.1928-3.2868	0.2819	124	0.2265	2624	X-RAY DIFFRACTION	100
3.2868-3.3928	0.2526	137	0.2251	2594	X-RAY DIFFRACTION	100
3.3928-3.5141	0.2842	134	0.2223	2606	X-RAY DIFFRACTION	99
3.5141-3.6547	0.2861	142	0.2201	2598	X-RAY DIFFRACTION	100
3.6547-3.821	0.2709	164	0.2106	2587	X-RAY DIFFRACTION	100
3.821-4.0224	0.2489	146	0.1872	2638	X-RAY DIFFRACTION	100
4.0224-4.2742	0.2157	159	0.1636	2621	X-RAY DIFFRACTION	100
4.2742-4.604	0.1814	128	0.1492	2673	X-RAY DIFFRACTION	100
4.604-5.0669	0.2309	134	0.1476	2651	X-RAY DIFFRACTION	99
5.0669-5.7991	0.1873	134	0.161	2698	X-RAY DIFFRACTION	100
5.7991-7.3025	0.2096	136	0.1819	2756	X-RAY DIFFRACTION	99
7.3025-49.1784	0.1973	147	0.1539	2905	X-RAY DIFFRACTION	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	0.6865	0.0598	-0.2663	1.2919	-0.1603	1.4103	-0.07	-0.1167	-0.1321	0.49	0.1857	-0.2523	0.0227	-0.0357	-0.1125	0.6187	0.1116	0.008	0.4683	-0.0377	0.5964	-29.1385	-57.724	50.0276
2	0.6838	-0.307	-0.2433	1.5064	0.5364	0.8638	-0.2455	-0.1	-0.2098	0.3498	0.1648	-0.0018	0.4481	0.0419	0.1367	0.4792	0.0823	0.0246	0.6031	0.0561	0.6068	-44.061	-64.4835	47.266
3	3.1364	-0.6396	-0.3612	2.2182	-0.106	1.2131	0.1965	0.6442	0.257	-0.4327	-0.1443	-0.6062	-0.0119	-0.0713	-0.0132	0.6694	-0.039	0.1934	0.7512	-0.0556	0.6073	-34.2232	-58.3206	10.2635
4	1.3126	-0.4493	0.1533	2.3395	-0.0105	0.9854	0.0068	0.125	0.2396	-0.0694	0.0394	-0.0433	-0.1534	-0.4202	-0.0558	0.5173	0.0344	0.1044	0.6777	0.0065	0.5959	-48.4371	-42.4137	27.0711
5	2.8651	-1.0526	-2.4571	1.5751	1.2797	4.4672	0.1066	-0.1697	0.0645	-0.0754	-0.0576	-0.0084	-0.2795	-0.3984	-0.0533	0.5062	0.1334	-0.0222	0.6176	0.0138	0.8101	-61.6621	-20.1779	47.6708
6	1.3576	-0.9331	0.675	1.4709	0.6387	1.76	-0.1989	0.5727	-0.6184	0.0365	-0.1998	0.2056	0.6799	-0.566	0.4062	0.5802	-0.1437	0.1265	0.9336	-0.0781	0.6789	-56.4399	-61.7204	21.8373
7	2.7357	0.4226	0.0351	3.1962	0.9508	1.8654	0.1821	0.1532	-0.1005	-0.6081	-0.3325	0.2585	0.3348	-0.9808	0.138	0.6707	-0.1421	-0.0083	1.0901	-0.0666	0.5857	-58.5231	-58.5579	14.2632
8	0.0035	-0.2982	-0.3937	1.4122	-0.7759	1.9968	-0.0055	-0.0235	0.1723	0.517	0.5034	-0.003	0.1201	-0.5027	-0.5313	0.7138	0.0923	0.1321	0.8552	-0.0234	0.6305	-40.7393	-56.0378	49.9642
9	0.8618	0.7354	1.0447	2.7135	0.033	1.6857	-0.0608	0.1233	0.305	0.8509	0.1927	0.3777	-0.3468	-1.2646	-0.1009	1.0786	0.1974	0.1379	1.1558	0.007	0.6989	-44.8128	-55.1411	60.3605
10	0.407	0.561	0.4118	2.0902	-0.7219	1.5545	-0.0974	0.2442	-0.4491	-0.1332	-0.0378	-0.654	-0.043	0.0475	0.1154	0.5021	-0.0302	0.1607	0.5436	-0.0392	0.6907	-27.4326	-57.0507	26.7203
11	6.1972	-3.4529	0.5728	5.4233	-2.6365	9.4418	-0.1684	-0.2168	-0.0158	-0.094	-0.4838	-1.1265	1.5163	-0.193	0.5396	1.3188	0.2404	0.2653	1.2284	-0.2511	1.7255	-5.4863	-75.8279	16.7803
12	3.3105	-0.0535	-0.2091	9.4003	2.9174	6.1399	0.3398	1.425	0.1365	-1.0138	-0.1136	-0.6738	0.1163	0.7937	-0.1778	0.975	0.1871	0.2048	0.9468	-0.2091	1.1677	-11.2246	-69.3782	15.1145

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	chain 'A' and (resid 1 through 356 )
2	X-RAY DIFFRACTION	2	chain 'A' and (resid 357 through 609 )
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 610 through 792 )
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 793 through 996 )
5	X-RAY DIFFRACTION	5	chain 'A' and (resid 997 through 1226 )
6	X-RAY DIFFRACTION	6	chain 'B' and (resid -20 through -11 )
7	X-RAY DIFFRACTION	7	chain 'B' and (resid -10 through -1 )
8	X-RAY DIFFRACTION	8	chain 'B' and (resid 0 through 19 )
9	X-RAY DIFFRACTION	9	chain 'C' and (resid -19 through -10 )
10	X-RAY DIFFRACTION	10	chain 'C' and (resid -9 through 5 )
11	X-RAY DIFFRACTION	11	chain 'C' and (resid 6 through 9 )
12	X-RAY DIFFRACTION	12	chain 'D' and (resid -9 through -1 )

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DNA chain , 2 types, 2 molecules CD

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Protein / RNA chain , 2 types, 2 molecules AB

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Non-polymers , 4 types, 23 molecules

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DNA chain , 2 types, 2 molecules CD

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