PDB-7r2a: Crystal structure of Ta_Cel5A Y200F variant, apoform

ID or keywords:

Entry

Database: PDB / ID: 7r2a

Title

Crystal structure of Ta_Cel5A Y200F variant, apoform

Components

EGI

Keywords

HYDROLASE / cellulase / GH5_5 family / GH-A clan / (a/b)8 fold / Thermoascus aurantiacus

Function / homology

glucan catabolic process / cellulase / Glycoside hydrolase, family 5 / Cellulase (glycosyl hydrolase family 5) / cellulase activity / Glycoside hydrolase superfamily / cellulase

Function and homology information

Biological species

Thermoascus aurantiacus ATCC 26904 (fungus)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT /

molecular replacement / Resolution: 1.3 Å

Authors

Dutoit, R.

Funding support

Belgium, 1items

Citation

Journal: To Be Published
Title: Crystal structure of Ta_Cel5A Y200F variant, apoform
Authors: Dutoit, R.

History

Deposition	Feb 4, 2022	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Feb 15, 2023	Provider: repository / Type: Initial release
Revision 1.1	Feb 7, 2024	Group: Data collection / Refinement description Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model
Revision 1.2	Nov 6, 2024	Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature / Item: _pdbx_entry_details.has_protein_modification

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	7r2a.cif.gz	271.3 KB	Display	PDBx/mmCIF format
PDB format	pdb7r2a.ent.gz	218.2 KB	Display	PDB format
PDBx/mmJSON format	7r2a.json.gz		Tree view	PDBx/mmJSON format
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Validation report

Summary document	7r2a_validation.pdf.gz	456.5 KB	Display	wwPDB validaton report
Full document	7r2a_full_validation.pdf.gz	459.6 KB	Display
Data in XML	7r2a_validation.xml.gz	33.1 KB	Display
Data in CIF	7r2a_validation.cif.gz	52.2 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/r2/7r2a ftp://data.pdbj.org/pub/pdb/validation_reports/r2/7r2a	HTTPS FTP

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Related structure data

Related structure data	1gzjS S: Starting model for refinement
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

Deposited unit

A: EGI

B: EGI

hetero molecules

68.4 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: EGI

hetero molecules

defined by author
Evidence: gel filtration, Ta_Cel5A Y200F elutes as a monomer using a Superdex75 column (V=120 mL)
34.3 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

B: EGI

hetero molecules

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

Unit cell

Length a, b, c (Å)	76.040, 84.780, 89.090
Angle α, β, γ (deg.)	90.000, 90.000, 90.000
Int Tables number	19
Space group name H-M	P2₁2₁2₁

#1: Protein	EGI / Endoglucanase Mass: 33713.328 Da / Num. of mol.: 2 / Mutation: Y200F Source method: isolated from a genetically manipulated source Source: (gene. exp.) Thermoascus aurantiacus ATCC 26904 (fungus) Gene: eg1 / Plasmid: pET22b / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q8TG26, cellulase
#2: Chemical	ChemComp-TRS / 2-AMINO-2-HYDROXYMETHYL-PROPANE-1,3-DIOL / TRIS BUFFER Mass: 122.143 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₄H₁₂NO₃ / Comment: pH buffer*YM
#3: Chemical	ChemComp-SO4 / SULFATE ION Mass: 96.063 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: SO₄
#4: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 892 / Source method: isolated from a natural source / Formula: H₂O
Has ligand of interest	N
Has protein modification	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: 2.13 Å³/Da / Density % sol: 42.24 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 6.5 Details: Reservoir solution: 0.1M Tris pH6.5, 1.6M ammonium sulfate Drop: 2 ul of 430 uM enzyme mixed with 2 ul of reservoir solution, 0.2 ul of microseeds

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

SOLEIL

/ Beamline: PROXIMA 1 / Wavelength: 0.9786 Å

Detector

Type: DECTRIS EIGER X 16M / Detector: PIXEL / Date: Jul 8, 2021

Radiation

Monochromator: Si (111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray

Radiation wavelength

Wavelength: 0.9786 Å / Relative weight: 1

Reflection

Resolution: 1.3→47.78 Å / Num. obs: 139574 / % possible obs: 99 % / Redundancy: 12.319 % / B_iso Wilson estimate: 15.4 Å² / CC_1/2: 0.999 / R_merge(I) obs: 0.086 / R_rim(I) all: 0.09 / Χ²: 0.869 / Net I/σ(I): 14.36 / Num. measured all: 1719356

Reflection shell

Diffraction-ID: 1

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Mean I/σ(I) obs	Num. measured obs	Num. possible	Num. unique obs	CC_1/2	R_rim(I) all	% possible all
1.3-1.34	11.34	1.77	1.49	102058	10327	9000	0.633	1.854	87.2
1.34-1.37	11.685	1.437	1.95	117329	10042	10041	0.767	1.503	100
1.37-1.41	11.601	1.115	2.52	113764	9810	9806	0.853	1.167	100
1.41-1.46	12.182	0.935	3.14	115501	9483	9481	0.904	0.977	100
1.46-1.5	11.759	0.709	4.07	108972	9267	9267	0.938	0.742	100
1.5-1.56	13.078	0.487	5.96	116589	8915	8915	0.971	0.507	100
1.56-1.62	13.143	0.357	7.48	113413	8629	8629	0.982	0.372	100
1.62-1.68	13.095	0.278	9.29	109013	8325	8325	0.987	0.29	100
1.68-1.76	13.05	0.244	11.15	103960	7967	7966	0.991	0.254	100
1.76-1.84	12.911	0.196	13.41	98743	7648	7648	0.992	0.204	100
1.84-1.94	12.67	0.164	15.39	91956	7258	7258	0.994	0.171	100
1.94-2.06	12.374	0.117	20.3	85792	6934	6933	0.996	0.122	100
2.06-2.2	11.857	0.093	23.94	76705	6472	6469	0.997	0.098	100
2.2-2.38	11.458	0.081	26.65	69492	6066	6065	0.998	0.085	100
2.38-2.6	12.358	0.068	31.73	68898	5575	5575	0.998	0.071	100
2.6-2.91	11.921	0.058	35.34	60559	5080	5080	0.998	0.061	100
2.91-3.36	13.206	0.051	41.73	59639	4516	4516	0.999	0.054	100
3.36-4.12	12.984	0.045	44.55	49755	3832	3832	0.999	0.047	100
4.12-5.82	12.679	0.042	45.49	38393	3028	3028	0.999	0.044	100
5.82-47.78	10.819	0.047	40.93	18825	1749	1740	0.999	0.05	99.5

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Phasing

Phasing

Method:

molecular replacement

Phasing MR

	Highest resolution	Lowest resolution
Rotation	3.59 Å	47.78 Å
Translation	3.59 Å	47.78 Å

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Processing

Software

Name	Version	Classification
XDS	20210323	data reduction
XSCALE	20210323	data scaling
PHASER	2.8.3	phasing
PHENIX	1.19.2	refinement
PDB_EXTRACT	3.27	data extraction

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 1GZJ

1gzj

Resolution: 1.3→47.78 Å / SU ML: 0.13 / Cross valid method: THROUGHOUT / σ(F): 1.36 / Phase error: 17.23 / Stereochemistry target values: ML

	R_factor	Num. reflection	% reflection
R_free	0.1668	6977	5 %
R_work	0.1527	132527	-
obs	0.1534	139504	99 %

Solvent computation

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

Displacement parameters

B_iso max: 57.14 Å² / B_iso mean: 20.3409 Å² / B_iso min: 10.28 Å²

Refinement step

Cycle: final / Resolution: 1.3→47.78 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4754	0	56	892	5702
B_iso mean	-	-	30.05	32.12	-
Num. residues	-	-	-	-	610

LS refinement shell

Refine-ID: X-RAY DIFFRACTION / R_factor R_free error: 0 / Total num. of bins used: 30

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Num. reflection all	% reflection obs (%)
1.3-1.32	0.3504	166	0.3355	3149	3315	72
1.32-1.33	0.2937	232	0.2736	4401	4633	100
1.33-1.35	0.2478	233	0.267	4427	4660	100
1.35-1.37	0.259	233	0.241	4417	4650	100
1.37-1.38	0.2731	232	0.2362	4406	4638	100
1.38-1.4	0.2346	231	0.2226	4370	4601	100
1.4-1.42	0.2376	233	0.2163	4426	4659	100
1.42-1.44	0.2337	232	0.2075	4429	4661	100
1.44-1.47	0.2304	234	0.2018	4444	4678	100
1.47-1.49	0.1998	233	0.1942	4434	4667	100
1.49-1.52	0.197	233	0.1676	4419	4652	100
1.52-1.54	0.1881	232	0.1519	4411	4643	100
1.54-1.57	0.169	233	0.1446	4413	4646	100
1.57-1.61	0.1568	234	0.1417	4459	4693	100
1.61-1.64	0.1568	234	0.1359	4435	4669	100
1.64-1.68	0.1558	233	0.1412	4436	4669	100
1.68-1.72	0.1754	235	0.1432	4459	4694	100
1.72-1.77	0.1774	233	0.1423	4420	4653	100
1.77-1.82	0.1669	234	0.1515	4445	4679	100
1.82-1.88	0.1662	235	0.1535	4459	4694	100
1.88-1.95	0.1925	234	0.1664	4459	4693	100
1.95-2.02	0.1815	236	0.1536	4474	4710	100
2.02-2.12	0.1593	235	0.1452	4468	4703	100
2.12-2.23	0.169	236	0.1449	4486	4722	100
2.23-2.37	0.153	236	0.1465	4481	4717	100
2.37-2.55	0.1445	237	0.1504	4509	4746	100
2.55-2.81	0.1567	237	0.1444	4499	4736	100
2.81-3.21	0.1699	239	0.1501	4544	4783	100
3.21-4.05	0.1414	242	0.1339	4597	4839	100
4.05-47.78	0.142	250	0.1352	4751	5001	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	1.076	-0.6532	-0.4523	1.4471	0.0502	1.9218	-0.0067	0.0263	-0.1301	0.0438	-0.0115	-0.0466	0.2363	0.0707	0.0435	0.1517	-0.0068	-0.0387	0.1137	-0.002	0.1552	5.4722	16.1712	9.8274
2	0.9062	-0.0424	-0.0586	1.1822	-0.3882	1.4295	0.0693	0.1559	-0.1449	-0.1871	-0.0562	0.0276	0.2325	0.0611	-0.0098	0.1701	0.0173	-0.0234	0.154	-0.0408	0.1587	7.3722	16.8469	-0.539
3	0.6278	-0.0298	0.0932	0.4393	-0.2379	0.7135	0.0029	-0.0015	-0.0228	0.0253	0.018	-0.0066	0.0109	-0.0099	-0.0205	0.1193	-0.0046	-0.0052	0.1166	-0.0045	0.1396	6.1887	26.0643	15.6154
4	1.1609	0.0039	-0.5144	0.9989	-0.1759	1.8074	-0.026	0.0734	-0.2693	-0.1717	-0.0011	0.1022	0.3249	-0.1066	0.0388	0.2325	-0.0196	-0.0432	0.1277	0.0034	0.2478	1.9094	5.3488	14.7707
5	1.4152	0.3173	-0.5595	1.7584	-0.0927	2.2025	-0.0979	0.1038	-0.1298	-0.0646	0.0385	-0.0672	0.3662	0.2091	0.0299	0.1532	0.0298	-0.0102	0.2006	-0.0125	0.1235	37.805	19.0498	34.232
6	1.7453	-0.097	-0.2403	1.2445	0.2868	1.6516	0.0007	-0.0635	-0.1771	0.12	0.0027	-0.0417	0.522	0.2823	0.0113	0.2379	0.069	-0.0135	0.1786	0.0018	0.1311	38.0813	14.6004	41.3138
7	0.3427	0.1604	0.0892	0.7128	-0.0992	1.5028	0.0058	-0.0351	-0.0377	0.0109	-0.0007	-0.0116	0.1637	0.0383	-0.0045	0.1305	-0.0009	0.0007	0.154	0.006	0.1326	29.8757	24.2825	44.1469
8	0.3968	0.0153	-0.0049	0.6534	0.2403	1.3351	-0.0274	0.02	0.0276	-0.0443	0.026	0.0045	-0.061	0.0911	-0.0004	0.118	-0.0055	-0.0018	0.1334	-0.0003	0.1382	31.7898	33.8104	33.966
9	0.4538	-0.3748	-0.0591	0.9684	0.1969	1.4426	0.0013	0.046	-0.0575	-0.155	-0.0156	0.0242	0.1025	0.1659	0.0153	0.1849	0.0092	0.0217	0.168	-0.0058	0.1363	35.3184	23.2833	18.1931
10	0.2667	-0.0234	0.091	0.7374	0.4008	1.2542	-0.0098	-0.0113	-0.0927	0.0341	0.0226	-0.1588	0.4978	0.483	-0.0022	0.2616	0.0975	0.0135	0.2285	-0.0117	0.16	42.1646	14.5596	24.9398

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	chain 'A' and (resid 1 through 20 )	A	1 - 20
2	X-RAY DIFFRACTION	2	chain 'A' and (resid 21 through 84 )	A	21 - 84
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 85 through 277 )	A	85 - 277
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 278 through 305 )	A	278 - 305
5	X-RAY DIFFRACTION	5	chain 'B' and (resid 1 through 20 )	B	1 - 20
6	X-RAY DIFFRACTION	6	chain 'B' and (resid 21 through 52 )	B	21 - 52
7	X-RAY DIFFRACTION	7	chain 'B' and (resid 53 through 105 )	B	53 - 105
8	X-RAY DIFFRACTION	8	chain 'B' and (resid 106 through 200 )	B	106 - 200
9	X-RAY DIFFRACTION	9	chain 'B' and (resid 201 through 246 )	B	201 - 246
10	X-RAY DIFFRACTION	10	chain 'B' and (resid 247 through 305 )	B	247 - 305

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