PDB-8g9a: Crystal structure of a resurrected ancestor (AncRNase) of the pan...

ID or keywords:

Entry

Database: PDB / ID: 8g9a

Title

Crystal structure of a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

Components

a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

Keywords

HYDROLASE / Ancestral sequence reconstruction / ASR / eosinophil-derived neurotoxin / EDN / RNase 2 / eosinophil cationic protein / ECP / RNase 3 / ribonuclease / transphosphorylase

Function / homology

ACETATE ION

Function and homology information

Biological species

synthetic construct (others)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.03 Å

Authors

Tran, T.T.Q. / Pham, N.T.H. / Calmettes, C. / Doucet, N.

Funding support

Canada, 5items

Organization	Grant number	Country
Natural Sciences and Engineering Research Council (NSERC, Canada)	RGPIN202204368	Canada
Natural Sciences and Engineering Research Council (NSERC, Canada)	RGPIN201706091	Canada
Natural Sciences and Engineering Research Council (NSERC, Canada)	CREATE511956	Canada
Fonds de Recherche du Quebec - Sante (FRQS)	251848	Canada
Fonds de Recherche du Quebec - Sante (FRQS)	281993	Canada

Citation

Journal: J.Biol.Chem. / Year: 2024
Title: Ancestral sequence reconstruction dissects structural and functional differences among eosinophil ribonucleases.
Authors: Tran, T.T.Q. / Narayanan, C. / Loes, A.N. / Click, T.H. / Pham, N.T.H. / Letourneau, M. / Harms, M.J. / Calmettes, C. / Agarwal, P.K. / Doucet, N.

History

Deposition	Feb 21, 2023	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Feb 28, 2024	Provider: repository / Type: Initial release
Revision 1.1	May 29, 2024	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.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	8g9a.cif.gz	334 KB	Display	PDBx/mmCIF format
PDB format	pdb8g9a.ent.gz	281.9 KB	Display	PDB format
PDBx/mmJSON format	8g9a.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	8g9a_validation.pdf.gz	494.9 KB	Display	wwPDB validaton report
Full document	8g9a_full_validation.pdf.gz	500.3 KB	Display
Data in XML	8g9a_validation.xml.gz	27.4 KB	Display
Data in CIF	8g9a_validation.cif.gz	39.4 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/g9/8g9a ftp://data.pdbj.org/pub/pdb/validation_reports/g9/8g9a	HTTPS FTP

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Related structure data

Related structure data	7ty1C 8f5xC C: citing same article (ref.)
Similar structure data	Similarity search - Function & homologyF&H Search Similarity search - HomologyF&H Search

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Links

PDB pages	PDBj / wwPDB / NCBI

Deposited unit

A: a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

B: a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

C: a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

D: a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

hetero molecules

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

1

A: a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

hetero molecules

defined by author
Evidence: gel filtration, 2D 1H-15N HSQC NMR titration experiments
16.4 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

B: a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

hetero molecules

defined by author
16.2 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

3

C: a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

hetero molecules

defined by author
15.9 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

4

D: a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families

hetero molecules

defined by author
16.3 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

Unit cell

Length a, b, c (Å)	170.290, 170.290, 170.290
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	197
Space group name H-M	I23

Components on special symmetry positions

ID	Model	Components
1	1	C-329- HOH

#1: Protein	a resurrected ancestor (AncRNase) of the pancreatic-type RNases 2 and 3 sub-families Mass: 15344.554 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli BL21(DE3) (bacteria) / References: EC: 3.1.27.5
#2: Chemical	... ChemComp-SO4 / SULFATE ION Mass: 96.063 Da / Num. of mol.: 24 / Source method: obtained synthetically / Formula: SO₄
#3: Chemical	ChemComp-ACT / ACETATE ION Mass: 59.044 Da / Num. of mol.: 14 / Source method: obtained synthetically / Formula: C₂H₃O₂
#4: Chemical	ChemComp-MPD / (4S)-2-METHYL-2,4-PENTANEDIOL Mass: 118.174 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₆H₁₄O₂ / Comment: precipitant*YM
#5: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 448 / Source method: isolated from a natural source / Formula: H₂O
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: 3.39 Å³/Da / Density % sol: 63.7 %
Crystal grow	Temperature: 298 K / Method: vapor diffusion, sitting drop / pH: 5.5 Details: 1.2 M Lithium sulfate, 0.1 M Sodium acetate:HCL pH 5.5 (cryoprotectant: 20% MPD)

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

CLSI

/ Beamline: 08B1-1 / Wavelength: 1.18067 Å

Detector

Type: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Mar 12, 2022

Radiation

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

Radiation wavelength

Wavelength: 1.18067 Å / Relative weight: 1

Reflection

Resolution: 2.03→45.52 Å / Num. obs: 52818 / % possible obs: 99.8 % / Redundancy: 38.9 % / CC_1/2: 1 / R_merge(I) obs: 0.089 / R_pim(I) all: 0.014 / R_rim(I) all: 0.09 / Net I/σ(I): 32.9

Reflection shell

Diffraction-ID: 1

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Mean I/σ(I) obs	Num. unique obs	CC_1/2	R_pim(I) all	R_rim(I) all	% possible all
2.03-2.1	36.4	1.874	2.8	4808	0.802	0.314	1.9	100
2.1-2.17	40.7	1.356	4.2	4769	0.903	0.215	1.373	100
2.17-2.26	39.9	0.936	6.1	4727	0.951	0.15	0.948	99.4
2.26-2.36	39.7	0.8	7.3	4738	0.968	0.128	0.81	99.5
2.36-2.48	39.9	0.495	11.2	4767	0.986	0.079	0.501	100
2.48-2.64	40.9	0.35	15.5	4807	0.993	0.055	0.354	100
2.64-2.84	39.7	0.225	23	4801	0.997	0.036	0.228	100
2.84-3.13	40	0.122	39.4	4797	0.999	0.02	0.124	100
3.13-3.58	39	0.071	62.5	4799	1	0.011	0.072	99.7
3.58-4.51	37.1	0.05	37.1	4834	1	0.008	0.051	99.6
4.51-45.52	35.3	0.037	100.8	4971	1	0.006	0.037	100

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Processing

Software

Name	Version	Classification
PHENIX	1.19.1-4122	refinement
XSCALE		data scaling
PDB_EXTRACT	4	data extraction
XDS		data reduction
PHASER	2.8.3	phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 2.03→45.51 Å / SU ML: 0.25 / Cross valid method: THROUGHOUT / σ(F): 1.35 / Phase error: 24.4 / Stereochemistry target values: ML

	R_factor	Num. reflection	% reflection
R_free	0.2054	1568	2.97 %
R_work	0.1911	-	-
obs	0.1915	52813	99.81 %

Solvent computation

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

Refinement step

Cycle: LAST / Resolution: 2.03→45.51 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4292	0	192	448	4932

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.015	4564
X-RAY DIFFRACTION	f_angle_d	1.455	6242
X-RAY DIFFRACTION	f_dihedral_angle_d	16.078	1664
X-RAY DIFFRACTION	f_chiral_restr	0.074	666
X-RAY DIFFRACTION	f_plane_restr	0.012	822

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 (%)
2.03-2.1	0.3349	142	0.2563	4666	X-RAY DIFFRACTION	100
2.1-2.17	0.2666	141	0.2302	4628	X-RAY DIFFRACTION	100
2.17-2.26	0.2964	136	0.2707	4591	X-RAY DIFFRACTION	100
2.26-2.36	0.2677	140	0.2354	4598	X-RAY DIFFRACTION	99
2.36-2.48	0.2335	140	0.2123	4627	X-RAY DIFFRACTION	100
2.48-2.64	0.2455	143	0.2357	4664	X-RAY DIFFRACTION	100
2.64-2.84	0.2572	141	0.2199	4660	X-RAY DIFFRACTION	100
2.85-3.13	0.2236	142	0.2143	4655	X-RAY DIFFRACTION	100
3.13-3.58	0.1954	148	0.1892	4651	X-RAY DIFFRACTION	100
3.58-4.51	0.1814	143	0.1537	4691	X-RAY DIFFRACTION	100
4.52-45.51	0.1663	152	0.1704	4814	X-RAY DIFFRACTION	100

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	0.6545	-0.6787	0.3305	0.9672	-0.5135	0.7849	-0.0791	-0.441	-0.2125	0.3555	0.0555	0.2069	-0.5213	-0.0265	-0	0.5077	0.0457	0.0531	0.3983	0.0467	0.3977	30.7823	26.0417	-43.2597
2	0.3576	0.0878	-0.2949	0.4253	-0.0947	0.3361	0.2014	0.1039	0.4372	-0.3135	-0.0544	-0.7381	-0.9055	0.7081	-0	0.5903	-0.076	0.0061	0.5029	0.0025	0.606	40.2249	14.9245	-46.6714
3	8.1977	-1.7651	-0.5164	1.4334	1.4426	2.0928	-0.0801	0.99	-0.8327	1.6526	-0.1881	1.6486	0.4402	-0.3234	0.2232	0.6051	-0.0019	0.2153	0.3433	0.1245	0.499	29.0388	14.8419	-40.9046
4	0.3624	-0.1418	-0.0585	0.2039	-0.1609	0.0402	0.0127	0.4887	0.1792	-0.2819	0.232	1.6895	-0.055	-0.4107	0.0001	0.5049	0.0183	0.02	0.3965	0.0159	0.6068	30.2401	15.3366	-52.0976
5	1.2168	-0.697	-0.1067	1.5822	-1.0892	3.328	-0.1816	0.1642	-0.073	0.0782	0.1276	0.178	-0.1164	-0.2955	0	0.3375	-0.0194	-0.0032	0.3073	0.0158	0.3573	31.4128	31.1506	-57.779
6	0.9909	-0.9527	-0.4363	1.2246	0.0868	0.3957	-0.081	-0.0628	-0.2218	-0.0696	-0.0532	0.0662	0.1913	0.2141	-0	0.365	-0.0088	-0.0195	0.2693	-0.006	0.3807	35.0698	10.5366	-51.6671
7	1.4138	-1.8345	0.7905	2.5707	-1.8607	2.0074	-0.2635	-0.123	-0.0435	0.3215	0.2412	0.4014	-0.62	-0.5603	0.0123	0.4832	0.0401	0.0347	0.304	0.035	0.346	30.0466	34.2121	-49.8436
8	1.439	0.8825	0.2615	0.8818	0.587	0.6662	0.0481	0.7444	-0.0508	-0.2504	-0.0208	-0.197	-0.0072	0.5343	0	0.4479	0.0428	0.0553	0.6271	-0.0148	0.4086	64.6653	29.429	-78.423
9	1.7245	0.5188	0.1255	0.3609	-0.6548	1.165	0.0452	0.3278	-0.3947	-0.0583	-0.1694	0.1166	0.2419	-0.0646	-0	0.4262	0.0544	-0.0068	0.4227	-0.0306	0.3772	58.8209	28.0698	-77.039
10	3.1692	-0.3439	-0.0317	0.9961	0.0775	1.4159	0.0653	0.4336	-0.4414	-0.1169	-0.0097	0.0319	0.1196	0.1787	0.0231	0.4022	0.0269	-0.0232	0.4074	-0.0754	0.3854	53.2346	24.5391	-75.72
11	2.0413	-0.1068	-0.3794	0.6449	-1.097	1.4251	0.0268	-0.0998	-0.4016	-0.1416	-0.033	-0.1013	0.1683	0.239	-0	0.4018	0.0501	0.0037	0.4361	0.0182	0.4251	60.6649	27.8432	-64.0287
12	0.6941	0.2195	-0.3856	0.2527	-0.2998	0.1037	-0.0634	0.3538	-0.2648	-0.0967	0.2739	0.05	0.2997	0.3254	-0	0.4221	0.0772	0.0591	0.5793	-0.0103	0.4475	80.0516	44.1754	-71.3612
13	0.6724	0.6715	-0.0657	1.2919	-0.0307	0.4957	0.4162	0.0334	-0.0305	-0.3425	-0.5208	-0.7887	-0.486	0.5956	-0.0004	0.5698	0.0767	0.1017	0.9438	0.0605	0.6371	91.0326	49.7302	-68.8248
14	1.4861	0.7271	0.0581	0.7688	0.1329	0.2544	0.0111	0.1815	0.0162	-0.1474	-0.0253	0.0438	-0.0454	0.22	-0	0.4081	-0.011	0.087	0.4819	0.0477	0.3754	72.2016	55.5188	-72.7002
15	1.9129	0.1171	-0.7543	1.4978	-0.6372	1.7614	0.0213	0.1608	0.0331	-0.1807	-0.2078	-0.3609	0.0927	0.7065	-0.3143	0.3881	0.0066	0.1129	0.6343	0.0956	0.4866	83.8687	57.5387	-72.8298
16	1.1821	-0.1318	1.268	1.2256	0.0756	1.1023	0.0959	-0.0196	-0.1642	-0.3329	-0.0515	0.1852	0.1403	0.143	0	0.4624	0.0171	0.0595	0.486	0.0108	0.3837	70.479	47.2096	-73.3124
17	1.4449	-1.5694	1.0275	2.0966	0.1484	2.2673	-0.0444	0.1681	-0.4742	0.0467	0.1658	0.1193	0.462	-0.1848	-0	0.409	0.0479	0.0094	0.42	-0.0188	0.4381	76.2104	25.7528	-53.0709
18	0.0723	0.1044	0.0266	0.7839	-1.2423	3.0359	-0.3869	-0.5141	-0.2783	-1.3393	0.773	-0.7263	1.7692	0.2187	0.1321	0.8454	0.0939	0.271	0.5718	-0.1217	0.7431	81.9126	18.4181	-53.6701
19	1.3405	0.793	-0.933	1.5417	0.9522	1.5801	-0.059	-0.0165	0.0339	0.2361	0.0177	-0.1651	0.0137	0.1757	-0	0.3041	0.0415	-0.0167	0.4004	0.0176	0.3498	74.4455	35.6317	-44.5884
20	0.0678	-0.0444	-0.1258	0.4969	-0.4358	0.0991	0.1981	-0.1475	0.077	-0.0718	-0.0375	-0.3112	0.1154	0.2733	0	0.3246	0.0311	-0.017	0.5137	0.0026	0.4148	84.5797	41.057	-40.946
21	0.9391	0.9123	-0.8641	1.3507	-0.1877	2.3011	-0.0954	-0.0715	-0.1355	0.0712	-0.027	-0.2659	0.2899	0.2912	-0.0138	0.2926	0.0979	0.0248	0.3608	0.0322	0.377	77.1473	28.9411	-46.236

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	(chain A and resid 0:17)
2	X-RAY DIFFRACTION	2	(chain A and resid 18:30)
3	X-RAY DIFFRACTION	3	(chain A and resid 31:37)
4	X-RAY DIFFRACTION	4	(chain A and resid 38:42)
5	X-RAY DIFFRACTION	5	(chain A and resid 43:82)
6	X-RAY DIFFRACTION	6	(chain A and resid 83:105)
7	X-RAY DIFFRACTION	7	(chain A and resid 106:132)
8	X-RAY DIFFRACTION	8	(chain B and resid 0:33)
9	X-RAY DIFFRACTION	9	(chain B and resid 34:52)
10	X-RAY DIFFRACTION	10	(chain B and resid 53:108)
11	X-RAY DIFFRACTION	11	(chain B and resid 109:132)
12	X-RAY DIFFRACTION	12	(chain C and resid 0:17)
13	X-RAY DIFFRACTION	13	(chain C and resid 18:40)
14	X-RAY DIFFRACTION	14	(chain C and resid 41:65)
15	X-RAY DIFFRACTION	15	(chain C and resid 66:109)
16	X-RAY DIFFRACTION	16	(chain C and resid 110:132)
17	X-RAY DIFFRACTION	17	(chain D and resid 0:27)
18	X-RAY DIFFRACTION	18	(chain D and resid 28:38)
19	X-RAY DIFFRACTION	19	(chain D and resid 39:61)
20	X-RAY DIFFRACTION	20	(chain D and resid 62:75)
21	X-RAY DIFFRACTION	21	(chain D and resid 76:132)

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Jul 12, 2017. Major update of PDB

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Yorodumi