PDB-5t0w: Crystal structure of the ancestral amino acid-binding protein Anc...

ID or keywords:

Entry

Database: PDB / ID: 5t0w

Title

Crystal structure of the ancestral amino acid-binding protein AncCDT-1, a precursor of cyclohexadienyl dehydratase

Components

AncCDT-1

Keywords

TRANSPORT PROTEIN / solute-binding protein / periplasmic binding protein / transport / amino acid

Function / homology

ARGININE

Function and homology information

Biological species

synthetic construct (others)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.59 Å

Authors

Clifton, B.E. / Carr, P.D. / Jackson, C.J.

Citation

Journal: To Be Published
Title: To be published
Authors: Clifton, B.E. / Kaczmarski, J.A. / Carr, P.D. / Jackson, C.J.

History

Deposition	Aug 16, 2016	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Sep 6, 2017	Provider: repository / Type: Initial release
Revision 1.1	Nov 1, 2017	Group: Author supporting evidence / Category: pdbx_struct_assembly_auth_evidence
Revision 1.2	Oct 4, 2023	Group: Data collection / Database references / Refinement description Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_ncs_dom_lim Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id / _struct_ncs_dom_lim.beg_label_comp_id / _struct_ncs_dom_lim.beg_label_seq_id / _struct_ncs_dom_lim.end_auth_comp_id / _struct_ncs_dom_lim.end_label_asym_id / _struct_ncs_dom_lim.end_label_comp_id / _struct_ncs_dom_lim.end_label_seq_id

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	5t0w.cif.gz	358.3 KB	Display	PDBx/mmCIF format
PDB format	pdb5t0w.ent.gz	295.1 KB	Display	PDB format
PDBx/mmJSON format	5t0w.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	5t0w_validation.pdf.gz	470.9 KB	Display	wwPDB validaton report
Full document	5t0w_full_validation.pdf.gz	483.7 KB	Display
Data in XML	5t0w_validation.xml.gz	32.9 KB	Display
Data in CIF	5t0w_validation.cif.gz	44.1 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/t0/5t0w ftp://data.pdbj.org/pub/pdb/validation_reports/t0/5t0w	HTTPS FTP

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Related structure data

Related structure data	4zv2S S: Starting model for refinement
Similar structure data	1iiw-d 2f7c-d 1v1f-d 3fms-d 4js7-d 1ii5-d 4bhp-d 4dz1-d 6q96-d 3b02-d Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Similarity search - Function & homologyF&H Search Similarity search - HomologyF&H Search

Related structure data

4zv2S

S: Starting model for refinement

Similar structure data

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Links

PDB pages	PDBj / wwPDB / NCBI
Related items in Molecule of the Month	#133 - Jan 2011 Nitric Oxide Synthase similarity (1)

Deposited unit

A: AncCDT-1

B: AncCDT-1

C: AncCDT-1

D: AncCDT-1

hetero molecules

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

1

A: AncCDT-1

hetero molecules

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

2

B: AncCDT-1

hetero molecules

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

3

C: AncCDT-1

hetero molecules

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

4

D: AncCDT-1

hetero molecules

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

Unit cell

Length a, b, c (Å)	47.030, 68.880, 318.630
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	19
Space group name H-M	P2₁2₁2₁

Noncrystallographic symmetry (NCS)

NCS domain:

ID	Ens-ID	Details
1	1	A
2	1	B
1	2	A
2	2	C
1	3	A
2	3	D
1	4	B
2	4	C
1	5	B
2	5	D
1	6	C
2	6	D

NCS domain segments:

Component-ID: _ / Beg auth comp-ID: SER / Beg label comp-ID: SER / End auth comp-ID: LYS / End label comp-ID: LYS / Refine code: _ / Auth seq-ID: 14 - 246 / Label seq-ID: 14 - 246

Dom-ID	Ens-ID	Auth asym-ID	Label asym-ID
1	1	A	A
2	1	B	B
1	2	A	A
2	2	C	C
1	3	A	A
2	3	D	D
1	4	B	B
2	4	C	C
1	5	B	B
2	5	D	D
1	6	C	C
2	6	D	D

NCS ensembles :

ID
1
2
3
4
5
6

Details

Monomer confirmed by size-exclusion chromatography

#1: Protein	AncCDT-1 Mass: 27779.801 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) synthetic construct (others) / Plasmid: pDOTS7 / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3)
#2: Chemical	ChemComp-ARG / ARGININE Type: L-peptide linking / Mass: 175.209 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Formula: C₆H₁₅N₄O₂
#3: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 21 / 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: 2.46 Å³/Da / Density % sol: 49.9 %
Crystal grow	Temperature: 291 K / Method: vapor diffusion, hanging drop / pH: 8.2 Details: A crystal grown in a hanging drop of 2 uL protein (18 mg/mL in 20 mM HEPES pH 7.5, 50 mM NaCl, 1 mM arginine) + 2 uL precipitant (0.2 M lithium sulfate, 0.1 M TRIS pH 8.2, 22% (w/v) PEG 3350) ...Details: A crystal grown in a hanging drop of 2 uL protein (18 mg/mL in 20 mM HEPES pH 7.5, 50 mM NaCl, 1 mM arginine) + 2 uL precipitant (0.2 M lithium sulfate, 0.1 M TRIS pH 8.2, 22% (w/v) PEG 3350) was improved by three rounds of serial microseeding; the crystals were crushed and serially diluted in the precipitant, and new hanging drops were prepared by mixing 2 uL protein and 2 uL microseed suspension.

Crystal

Density Matthews: 2.46 Å³/Da / Density % sol: 49.9 %

Crystal grow

Temperature: 291 K / Method: vapor diffusion, hanging drop / pH: 8.2
Details: A crystal grown in a hanging drop of 2 uL protein (18 mg/mL in 20 mM HEPES pH 7.5, 50 mM NaCl, 1 mM arginine) + 2 uL precipitant (0.2 M lithium sulfate, 0.1 M TRIS pH 8.2, 22% (w/v) PEG 3350) ...

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Data collection

Diffraction	Mean temperature: 100 K
Diffraction source	Source: SYNCHROTRON / Site: Australian Synchrotron / Beamline: MX1 / Wavelength: 0.9501 Å
Detector	Type: ADSC QUANTUM 210r / Detector: CCD / Date: Oct 29, 2015
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 0.9501 Å / Relative weight: 1
Reflection	Resolution: 2.59→33.66 Å / Num. obs: 33106 / % possible obs: 99.4 % / Redundancy: 6.8 % / B_iso Wilson estimate: 29.2 Å² / CC_1/2: 0.993 / R_merge(I) obs: 0.158 / Net I/σ(I): 8.2
Reflection shell	Resolution: 2.59→2.71 Å / Redundancy: 6.9 % / R_merge(I) obs: 0.775 / Mean I/σ(I) obs: 2.4 / CC_1/2: 0.755 / % possible all: 99.3

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Processing

Software

Name	Version	Classification
REFMAC	5.7.0032	refinement
iMOSFLM	1.0.7	data reduction
Aimless	0.1.29	data scaling
PHASER	2.5.2	phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 4ZV2

4zv2

Resolution: 2.59→33.19 Å / Cor.coef. F_o:F_c: 0.888 / Cor.coef. F_o:F_c free: 0.863 / SU B: 31.219 / SU ML: 0.311 / Cross valid method: THROUGHOUT / ESU R: 0.971 / ESU R Free: 0.359 / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS

	R_factor	Num. reflection	% reflection	Selection details
R_free	0.28733	1686	5.1 %	RANDOM
R_work	0.25348	-	-	-
obs	0.25518	31387	98.87 %	-

Solvent computation

Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å

Displacement parameters

B_iso mean: 43.419 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	2.35 Å²	-0 Å²	-0 Å²
2-	-	0.55 Å²	0 Å²
3-	-	-	-2.9 Å²

Refinement step

Cycle: LAST / Resolution: 2.59→33.19 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	6937	0	48	21	7006

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.011	0.02	7133
X-RAY DIFFRACTION	r_bond_other_d	0.006	0.02	6702
X-RAY DIFFRACTION	r_angle_refined_deg	1.372	1.958	9687
X-RAY DIFFRACTION	r_angle_other_deg	1.195	3	15392
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	6.038	5	902
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	31.911	24.966	298
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	18.623	15	1151
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	13.699	15	26
X-RAY DIFFRACTION	r_chiral_restr	0.077	0.2	1106
X-RAY DIFFRACTION	r_gen_planes_refined	0.007	0.021	8063
X-RAY DIFFRACTION	r_gen_planes_other	0.006	0.02	1551
X-RAY DIFFRACTION	r_nbd_refined
X-RAY DIFFRACTION	r_nbd_other
X-RAY DIFFRACTION	r_nbtor_refined
X-RAY DIFFRACTION	r_nbtor_other
X-RAY DIFFRACTION	r_xyhbond_nbd_refined
X-RAY DIFFRACTION	r_xyhbond_nbd_other
X-RAY DIFFRACTION	r_metal_ion_refined
X-RAY DIFFRACTION	r_metal_ion_other
X-RAY DIFFRACTION	r_symmetry_vdw_refined
X-RAY DIFFRACTION	r_symmetry_vdw_other
X-RAY DIFFRACTION	r_symmetry_hbond_refined
X-RAY DIFFRACTION	r_symmetry_hbond_other
X-RAY DIFFRACTION	r_symmetry_metal_ion_refined
X-RAY DIFFRACTION	r_symmetry_metal_ion_other
X-RAY DIFFRACTION	r_mcbond_it	1.045	1.64	3644
X-RAY DIFFRACTION	r_mcbond_other	1.046	1.64	3643
X-RAY DIFFRACTION	r_mcangle_it	1.86	2.451	4531
X-RAY DIFFRACTION	r_mcangle_other	1.859	2.451	4532
X-RAY DIFFRACTION	r_scbond_it	0.836	1.671	3489
X-RAY DIFFRACTION	r_scbond_other	0.836	1.671	3489
X-RAY DIFFRACTION	r_scangle_it
X-RAY DIFFRACTION	r_scangle_other	1.5	2.488	5156
X-RAY DIFFRACTION	r_long_range_B_refined	3.106	12.934	8033
X-RAY DIFFRACTION	r_long_range_B_other	3.103	12.933	8033
X-RAY DIFFRACTION	r_rigid_bond_restr
X-RAY DIFFRACTION	r_sphericity_free
X-RAY DIFFRACTION	r_sphericity_bonded

Refine LS restraints NCS

Refine-ID: X-RAY DIFFRACTION / Type: interatomic distance / Weight position: 0.05

Ens-ID	Dom-ID	Auth asym-ID	Number	Rms dev position (Å)
1	1	A	12532	0.11
1	2	B	12532	0.11
2	1	A	12031	0.11
2	2	C	12031	0.11
3	1	A	12237	0.11
3	2	D	12237	0.11
4	1	B	12329	0.09
4	2	C	12329	0.09
5	1	B	12559	0.1
5	2	D	12559	0.1
6	1	C	12212	0.08
6	2	D	12212	0.08

LS refinement shell

Resolution: 2.59→2.657 Å / Total num. of bins used: 20

	R_factor	Num. reflection	% reflection
R_free	0.374	128	-
R_work	0.348	2254	-
obs	-	-	98.59 %

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	1.2674	-0.0654	0.2881	3.2775	-0.9816	2.0225	-0.0616	0.0998	0.0991	-0.3325	0.0704	-0.0312	0.3444	-0.1992	-0.0088	0.2094	-0.0267	-0.0208	0.2622	-0.0222	0.2346	-9.8205	4.0655	-6.6871
2	2.7257	-2.1284	-0.3335	11.4797	-1.6557	1.2742	0.2535	0.1715	0.1362	0.2029	-0.0177	-0.0233	-0.4136	-0.1181	-0.2358	0.2015	-0.0162	-0.0064	0.2673	0.0573	0.2562	-14.7166	25.6374	-15.512
3	4.605	-1.5328	1.2992	2.0267	-2.8707	4.2894	0.1419	0.1562	-0.4493	-0.5036	-0.09	0.0829	0.7625	0.0415	-0.0519	0.2052	0.0094	-0.0755	0.2256	-0.1131	0.2647	-9.3957	14.9908	-21.6344
4	1.3929	0.9018	-0.1951	2.0913	-1.1043	2.542	-0.1314	0.2363	0.1773	-0.0788	0.0411	-0.2087	-0.0482	-0.0008	0.0903	0.1382	0.0337	0.0216	0.2415	-0.0099	0.2793	-1.6185	6.8641	-1.6557
5	3.5111	0.7703	-2.3019	2.2613	-0.6855	4.2883	0.2076	-0.2529	0.1438	-0.215	-0.0249	0.023	-0.2097	0.143	-0.1826	0.2055	-0.0128	-0.0679	0.2117	-0.0412	0.1478	-36.1053	36.8769	-73.8163
6	9.1556	0.9314	-1.3212	1.0587	-1.9946	7.084	-0.0911	-0.3331	-0.3987	-0.149	0.1804	0.5532	0.1366	-0.9906	-0.0893	0.0367	0.0459	-0.1186	0.3947	-0.0152	0.687	-58.8463	37.5758	-61.7279
7	1.4302	-1.2721	-2.632	6.8744	1.8282	5.2507	-0.0191	-0.1018	-0.1338	0.0236	-0.0353	-0.0884	0.1323	-0.1161	0.0544	0.1226	-0.0465	-0.1219	0.3763	0.078	0.2703	-48.084	32.5284	-57.3263
8	3.2057	-0.144	-2.1443	3.7134	-0.1559	3.5747	-0.0893	-0.0519	-0.1991	-0.4457	0.0393	0.1239	0.4012	-0.3913	0.0501	0.3091	-0.0349	-0.0951	0.2582	-0.0068	0.1672	-37.5844	28.4126	-80.4919
9	2.8077	-0.7096	2.0174	4.495	3.0916	5.6879	-0.2108	0.1587	0.0126	-0.5465	0.6316	-0.5368	0.1187	0.8514	-0.4208	0.6148	0.0112	0.0312	0.1617	-0.0902	0.1422	-9.1238	5.2359	-45.3885
10	4.217	1.3448	3.6886	3.0576	4.3474	7.0779	-0.0777	0.0061	-0.2349	0.324	-0.0972	0.2015	0.3246	-0.1999	0.1749	0.6001	-0.0436	-0.0167	0.0838	-0.0219	0.0638	-21.7664	-3.8516	-45.5844
11	7.6997	4.3859	-0.1792	5.9397	3.7176	8.0743	0.1016	0.675	0.1872	0.6894	-0.2061	0.5726	0.6403	-0.635	0.1046	0.6843	-0.1189	0.0334	0.16	-0.0605	0.1282	-28.3989	-13.8379	-58.1054
12	1.5073	0.2832	1.146	2.1066	3.4689	6.0539	-0.2053	-0.1346	0.2214	0.1594	-0.1947	0.188	0.2713	-0.3996	0.4	0.5578	-0.0611	-0.0589	0.0932	0.0055	0.1453	-23.3189	2.374	-48.0627
13	2.0571	0.1777	1.8104	2.9433	0.6723	4.6841	0.1988	0.1261	-0.1431	-0.1029	-0.1825	-0.0038	-0.0773	-0.1647	-0.0163	0.1751	0.002	-0.0175	0.248	0.0243	0.2136	-32.9987	36.3928	-34.5683
14	0.0225	0.4271	-0.0662	20.3715	-3.9445	0.8067	0.0279	-0.0721	0.0497	0.1128	-0.2416	-0.2722	-0.0626	0.0009	0.2137	0.2581	-0.1592	-0.1772	0.6315	-0.1135	0.5917	-16.0574	58.5738	-16.913
15	2.0069	-0.8209	2.1304	3.8371	-0.2156	8.2883	-0.1547	0.2812	0.3776	0.0259	-0.3752	-0.4222	-0.5989	0.9535	0.5299	0.23	-0.0967	-0.0482	0.1965	0.1451	0.2409	-20.436	45.8184	-19.6091
16	2.8449	0.0053	1.1037	3.4588	1.5847	4.7756	-0.055	0.1438	0.1825	-0.3185	-0.2646	0.193	-0.6904	-0.3729	0.3195	0.2548	0.1077	-0.0309	0.2441	0.0375	0.137	-37.2691	45.3331	-36.6987

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	A	14 - 125
2	X-RAY DIFFRACTION	2	A	126 - 158
3	X-RAY DIFFRACTION	3	A	159 - 195
4	X-RAY DIFFRACTION	4	A	196 - 246
5	X-RAY DIFFRACTION	5	B	14 - 119
6	X-RAY DIFFRACTION	6	B	120 - 159
7	X-RAY DIFFRACTION	7	B	160 - 201
8	X-RAY DIFFRACTION	8	B	204 - 246
9	X-RAY DIFFRACTION	9	C	14 - 83
10	X-RAY DIFFRACTION	10	C	84 - 118
11	X-RAY DIFFRACTION	11	C	119 - 163
12	X-RAY DIFFRACTION	12	C	164 - 246
13	X-RAY DIFFRACTION	13	D	13 - 118
14	X-RAY DIFFRACTION	14	D	119 - 133
15	X-RAY DIFFRACTION	15	D	134 - 186
16	X-RAY DIFFRACTION	16	D	187 - 246

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Yorodumi