PDB-7r28: Crystal structure of Ta_Cel5A E133Q Y200F variant, apoform

ID or keywords:

Entry

Database: PDB / ID: 7r28

Title

Crystal structure of Ta_Cel5A E133Q Y200F variant, apoform

Components

EGI

Keywords

HYDROLASE /

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

Function / homology

Glycoside hydrolase, family 5 / Cellulase (glycosyl hydrolase family 5) /

cellulase /

cellulase activity / cellulose catabolic process / 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: 1.22 Å

Authors

Dutoit, R.

Funding support

Belgium, 1items

Citation

Journal: To Be Published
Title: Crystal structure of Ta_Cel5A E133Q 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

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	7r28.cif.gz	273.2 KB	Display	PDBx/mmCIF format
PDB format	pdb7r28.ent.gz	220.2 KB	Display	PDB format
PDBx/mmJSON format	7r28.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/r2/7r28 ftp://data.pdbj.org/pub/pdb/validation_reports/r2/7r28	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

68.1 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 E133Q Y20F elutes as a monomer using a Superdex 200 (V=120mL)
34 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)	76.030, 85.100, 89.510
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: 33712.344 Da / Num. of mol.: 2 / Mutation: E133Q Y200F Source method: isolated from a genetically manipulated source Source: (gene. exp.) Thermoascus aurantiacus ATCC 26904 (fungus) Gene: eg1 / Plasmid: pET30b / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q8TG26, cellulase
#2: Chemical	ChemComp-SO4 / SULFATE ION / Sulfate Mass: 96.063 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: SO₄
#3: Water	ChemComp-HOH / water / Water Mass: 18.015 Da / Num. of mol.: 1007 / 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: 2.15 Å³/Da / Density % sol: 42.72 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 7 Details: Reservoir solution: 0.1M Tris pH7.0, 1.9M ammonium sulfate Drop: 2 ul of 465 uM enzyme mixed with 2 ul of reservoir solution and 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: Apr 2, 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: 1.22→42.55 Å / Num. obs: 170353 / % possible obs: 99.8 % / Redundancy: 13.211 % / B_iso Wilson estimate: 15.19 Å² / CC_1/2: 1 / R_merge(I) obs: 0.059 / R_rim(I) all: 0.062 / Χ²: 0.881 / Net I/σ(I): 20.41 / Num. measured all: 2250559 / Scaling rejects: 2

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
1.22-1.26	12.055	1.297	1.72	145954	12490	12107	0.762	1.355	96.9
1.26-1.29	13.316	1.095	2.33	162697	12218	12218	0.868	1.139	100
1.29-1.33	13.147	0.883	2.96	155835	11854	11853	0.916	0.919	100
1.33-1.37	12.677	0.683	3.75	146390	11549	11548	0.944	0.712	100
1.37-1.41	12.899	0.511	5.1	143955	11160	11160	0.97	0.532	100
1.41-1.46	12.956	0.373	6.77	140961	10880	10880	0.982	0.388	100
1.46-1.52	13.803	0.275	9.38	144169	10445	10445	0.989	0.286	100
1.52-1.58	13.752	0.212	11.95	138329	10059	10059	0.992	0.22	100
1.58-1.65	13.664	0.164	14.95	132474	9695	9695	0.995	0.17	100
1.65-1.73	13.476	0.132	18.16	124536	9241	9241	0.996	0.137	100
1.73-1.82	12.804	0.103	22.6	112789	8810	8809	0.997	0.107	100
1.82-1.94	13.325	0.084	28.24	111652	8379	8379	0.998	0.088	100
1.94-2.07	13.348	0.064	36.45	104809	7852	7852	0.999	0.067	100
2.07-2.23	14.167	0.055	43.61	104325	7364	7364	0.999	0.057	100
2.23-2.45	13.946	0.05	47.66	94149	6751	6751	0.999	0.052	100
2.45-2.74	13.655	0.044	52.25	83924	6146	6146	0.999	0.046	100
2.74-3.16	12.703	0.038	57.28	69471	5469	5469	0.999	0.039	100
3.16-3.87	12.509	0.034	63.7	58029	4639	4639	1	0.035	100
3.87-5.47	13.581	0.03	70.08	49406	3638	3638	1	0.031	100
5.47-42.55	12.717	0.033	66.22	26705	2116	2100	0.999	0.035	99.2

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Phasing

Phasing

Method:

molecular replacement

Phasing MR

	Highest resolution	Lowest resolution
Rotation	3.6 Å	47.9 Å
Translation	3.6 Å	47.9 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: 1.22→42.55 Å / SU ML: 0.15 / Cross valid method: THROUGHOUT / σ(F): 1.36 / Phase error: 16.53 / Stereochemistry target values: ML

	R_factor	Num. reflection	% reflection
R_free	0.166	8513	5 %
R_work	0.1567	161769	-
obs	0.1572	170282	99.72 %

Solvent computation

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

Displacement parameters

B_iso max: 65.75 Å² / B_iso mean: 19.6559 Å² / B_iso min: 9.91 Å²

Refinement step

Cycle: final / Resolution: 1.22→42.55 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4749	0	35	1007	5791
B_iso mean	-	-	32.1	31.17	-
Num. residues	-	-	-	-	609

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 (%)
1.22-1.24	0.3929	261	0.4042	4967	5228	93
1.24-1.25	0.3671	281	0.324	5344	5625	100
1.25-1.27	0.3202	282	0.2942	5349	5631	100
1.27-1.28	0.2984	280	0.2786	5331	5611	100
1.28-1.3	0.2572	283	0.2596	5364	5647	100
1.3-1.32	0.2579	282	0.244	5361	5643	100
1.32-1.34	0.2295	282	0.218	5346	5628	100
1.34-1.36	0.2413	284	0.2056	5395	5679	100
1.36-1.38	0.2026	281	0.1904	5336	5617	100
1.38-1.4	0.1903	282	0.178	5363	5645	100
1.4-1.43	0.1947	281	0.1833	5358	5639	100
1.43-1.45	0.196	283	0.1852	5380	5663	100
1.45-1.48	0.2072	281	0.1939	5351	5632	100
1.48-1.51	0.197	283	0.1956	5377	5660	100
1.51-1.54	0.1908	286	0.1655	5425	5711	100
1.54-1.58	0.1633	282	0.1555	5350	5632	100
1.58-1.62	0.1721	282	0.1498	5368	5650	100
1.62-1.66	0.1514	284	0.1404	5393	5677	100
1.66-1.71	0.1806	284	0.1472	5397	5681	100
1.71-1.77	0.1617	285	0.144	5415	5700	100
1.77-1.83	0.1649	283	0.1498	5378	5661	100
1.83-1.9	0.161	287	0.1559	5452	5739	100
1.9-1.99	0.1661	285	0.1483	5411	5696	100
1.99-2.09	0.147	284	0.1421	5406	5690	100
2.09-2.22	0.1479	287	0.1446	5435	5722	100
2.22-2.4	0.1648	286	0.1464	5442	5728	100
2.4-2.64	0.1318	289	0.1514	5486	5775	100
2.64-3.02	0.1746	290	0.1499	5506	5796	100
3.02-3.8	0.1468	291	0.1438	5543	5834	100
3.8-42.55	0.141	302	0.1348	5740	6042	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	0.0424	0.0685	0.0823	0.0959	0.1761	0.2439	0.0197	-0.0693	-0.1831	-0.0433	-0.0225	-0.0278	0.156	0.0816	-0.0002	0.1403	0.0006	-0.0198	0.1287	0.0003	0.1677	-5.4133	16.4019	-9.8308
2	0.1255	0.1211	0.0075	0.4723	0.3687	0.5897	0.0667	-0.1032	-0.2327	0.1483	-0.0623	-0.0424	0.2158	-0.0369	-0.1226	0.1768	0.0075	-0.023	0.1534	0.05	0.2202	-4.8531	12.1189	-2.8565
3	0.0952	0.1305	0.103	0.2137	0.0902	0.2286	0.0599	-0.2056	-0.0993	0.1824	-0.0569	0.0045	0.085	-0.0652	-0	0.1546	-0.0133	-0.0072	0.2011	0.0342	0.1596	-9.6245	21.5659	3.6383
4	0.165	0.2227	-0.0114	0.3649	0.0656	0.2687	0.0016	-0.0911	0.0145	0.02	-0.0006	0.0625	0.0471	-0.05	-0	0.115	0.011	-0.0024	0.1421	-0.001	0.1434	-11.2988	28.584	-5.2808
5	0.4904	0.0892	0.0693	0.2361	0.0416	0.2139	-0.0216	-0.0039	0.0077	-0.0289	0.0311	0.0296	-0.0332	0.0258	-0	0.1188	0.0016	-0.0057	0.1197	0.0005	0.1349	-7.205	29.1629	-16.7221
6	0.0301	-0.0055	-0.0015	0.0131	0.0093	-0	-0.0179	0.1476	-0.0103	-0.1891	-0.0043	0.0394	-0.0907	0.0548	0	0.1747	-0.0094	0.0047	0.1593	-0.0132	0.1476	-1.8435	27.0066	-29.1816
7	0.1884	-0.0786	0.1392	0.0412	-0.043	0.0947	0.0423	0.071	-0.0623	-0.0504	-0.0282	0.0205	-0.0122	0.0771	0	0.1401	0.0039	0.0008	0.1213	-0.0141	0.1553	-2.1715	21.2411	-20.4543
8	0.2718	-0.0444	0.0254	0.1968	-0.0537	0.3147	0.0755	0.0338	-0.1286	0.0047	-0.0192	-0.0207	0.0482	0.2128	-0.0052	0.1364	0.0227	-0.008	0.1428	-0.0314	0.1629	1.1353	16.146	-22.679
9	0.1338	0.0306	0.0304	0.07	0.126	0.283	-0.0329	-0.0345	-0.2997	0.1473	0.0087	0.0015	0.3441	0.066	0.0186	0.2149	0.0344	-0.0416	0.1275	-0.0055	0.2536	-1.757	5.5124	-14.8353
10	0.1958	-0.0404	0.1018	0.0441	-0.0793	0.2359	-0.0331	-0.0255	-0.1153	0.0522	-0.0005	0.0515	0.2636	-0.1619	0.0034	0.1312	-0.0124	0.0113	0.186	0.0047	0.1371	-38.2165	19.276	-34.2546
11	0.1286	-0.0283	0.0893	0.213	-0.3024	0.3771	0.0213	0.0297	-0.0859	-0.0914	0.0046	-0.003	0.2818	-0.1783	0.014	0.1793	-0.045	-0.0045	0.1835	-0.0091	0.1396	-38.5786	14.5542	-41.499
12	0.4315	-0.0249	0.0986	0.3792	-0.0132	0.9011	-0.0145	0.0429	0.0064	0.0125	0.0151	-0.0176	0.0004	-0.0213	0	0.1039	-0.0012	-0.0025	0.1242	0.0009	0.1182	-30.4518	29.9825	-38.7541
13	0.3127	0.0216	0.2299	0.4147	-0.1363	1.0136	0.0406	-0.0587	-0.0418	0.0899	-0.0242	0.0504	0.1784	-0.2257	-0.0007	0.1734	-0.0335	0.0185	0.1647	0.0078	0.1281	-39.1062	20.4513	-23.284

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 2 through 20 )	A	2 - 20
2	X-RAY DIFFRACTION	2	chain 'A' and (resid 21 through 52 )	A	21 - 52
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 53 through 84 )	A	53 - 84
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 85 through 138 )	A	85 - 138
5	X-RAY DIFFRACTION	5	chain 'A' and (resid 139 through 216 )	A	139 - 216
6	X-RAY DIFFRACTION	6	chain 'A' and (resid 217 through 231 )	A	217 - 231
7	X-RAY DIFFRACTION	7	chain 'A' and (resid 232 through 246 )	A	232 - 246
8	X-RAY DIFFRACTION	8	chain 'A' and (resid 247 through 277 )	A	247 - 277
9	X-RAY DIFFRACTION	9	chain 'A' and (resid 278 through 305 )	A	278 - 305
10	X-RAY DIFFRACTION	10	chain 'B' and (resid 1 through 20 )	B	1 - 20
11	X-RAY DIFFRACTION	11	chain 'B' and (resid 21 through 52 )	B	21 - 52
12	X-RAY DIFFRACTION	12	chain 'B' and (resid 53 through 185 )	B	53 - 185
13	X-RAY DIFFRACTION	13	chain 'B' and (resid 186 through 305 )	B	186 - 305

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