PDB-7r2d: Crystal structure of TaCel5A E133A variant in complex with cellop...

ID or keywords:

Entry

Database: PDB / ID: 7r2d

Title

Crystal structure of TaCel5A E133A variant in complex with cellopentaose

Components

EGI

Keywords

HYDROLASE /

Glycoside hydrolase /

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

Function / homology

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

cellulase /

cellulase activity / cellulose catabolic process / Glycoside hydrolase superfamily / beta-cellopentaose /

cellulase

Function and homology information

Biological species

Thermoascus aurantiacus ATCC 26904 (fungus)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT /

molecular replacement / Resolution: 1.61 Å

Authors

Dutoit, R.

Funding support

Belgium, 1items

Citation

Journal: To Be Published
Title: Crystal structure of TaCel5A E133A variant in complex with cellopentaose
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	7r2d.cif.gz	273.9 KB	Display	PDBx/mmCIF format
PDB format	pdb7r2d.ent.gz	221.7 KB	Display	PDB format
PDBx/mmJSON format	7r2d.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/r2/7r2d ftp://data.pdbj.org/pub/pdb/validation_reports/r2/7r2d	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
Related items in Molecule of the Month	#254 - Feb 2021 Cellulose Synthase similarity (1)

Deposited unit

A: EGI

B: EGI

hetero molecules

69.7 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 E133A elutes as a monomer on a Superdex 75 column (V=120 ml)
34.9 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

B: EGI

hetero molecules

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

Unit cell

Length a, b, c (Å)	75.200, 84.940, 89.040
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: 33671.293 Da / Num. of mol.: 2 / Mutation: E133A
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: Polysaccharide

beta-D-glucopyranose-(1-4)-beta-D-glucopyranose-(1-4)-beta-D-glucopyranose-(1-4)-beta-D- ...

Type: oligosaccharide

, Oligosaccharide

/ Class: Metabolism

/ Mass: 828.719 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: oligosaccharide / References: beta-cellopentaose

Descriptor	Type	Program
DGlcpb1-4DGlcpb1-4DGlcpb1-4DGlcpb1-4DGlcpb1-ROH	Glycam Condensed Sequence	GMML 1.0
WURCS=2.0/1,5,4/[a2122h-1b_1-5]/1-1-1-1-1/a4-b1_b4-c1_c4-d1_d4-e1	WURCS	PDB2Glycan 1.1.0
[][b-D-Glcp]{[(4+1)][b-D-Glcp]{[(4+1)][b-D-Glcp]{[(4+1)][b-D-Glcp]{[(4+1)][b-D-Glcp]{}}}}}	LINUCS	PDB-CARE

#3: Chemical

ChemComp-SO4 / SULFATE ION /

Sulfate

Mass: 96.063 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: SO₄

#4: Water

ChemComp-HOH / water /

Water

Mass: 18.015 Da / Num. of mol.: 838 / Source method: isolated from a natural source / Formula: H₂O

Has ligand of interest

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: 2.11 Å³/Da / Density % sol: 41.74 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 8 Details: Reservoir solution: 0.1M Tris pH8.0, 1.9M ammonium sulfate Drop: 2 ul of 430 uM enzyme, 2 ul of reservoir solution, 0.2 ul of microseeds, 0.2 ul of 10 mM cellopentaose

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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: 1.61→42.47 Å / Num. obs: 73962 / % possible obs: 99.6 % / Redundancy: 13.365 % / B_iso Wilson estimate: 20.35 Å² / CC_1/2: 0.999 / R_merge(I) obs: 0.118 / R_rim(I) all: 0.123 / Χ²: 0.88 / Net I/σ(I): 13.39 / Num. measured all: 988515 / Scaling rejects: 127

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
1.61-1.65	11.442	1.449	1.79	58767	5407	5136	0.741	1.516	95
1.65-1.7	13.517	1.221	2.46	71856	5316	5316	0.864	1.269	100
1.7-1.75	13.119	0.975	2.99	67548	5150	5149	0.903	1.014	100
1.75-1.8	14.125	0.755	4.01	70627	5000	5000	0.949	0.784	100
1.8-1.86	14.057	0.618	4.84	68162	4849	4849	0.96	0.642	100
1.86-1.93	13.96	0.535	5.84	65641	4703	4702	0.971	0.555	100
1.93-2	13.846	0.362	7.79	62655	4525	4525	0.986	0.376	100
2-2.08	13.684	0.275	9.87	60045	4388	4388	0.99	0.286	100
2.08-2.17	13.355	0.228	11.65	56023	4195	4195	0.992	0.237	100
2.17-2.28	12.535	0.217	12.62	50480	4033	4027	0.991	0.226	99.9
2.28-2.4	13.494	0.159	15.85	51600	3824	3824	0.995	0.166	100
2.4-2.55	13.321	0.138	17.94	48317	3627	3627	0.996	0.144	100
2.55-2.72	14.232	0.115	21.9	48646	3418	3418	0.997	0.119	100
2.72-2.94	13.987	0.099	25.09	44772	3201	3201	0.998	0.102	100
2.94-3.22	13.66	0.084	28.71	40365	2955	2955	0.998	0.088	100
3.22-3.6	13.131	0.073	31.86	35243	2684	2684	0.998	0.076	100
3.6-4.16	11.975	0.063	34.2	28524	2384	2382	0.998	0.066	99.9
4.16-5.1	12.544	0.056	36.99	25539	2036	2036	0.999	0.058	100
5.1-7.21	13.771	0.052	37.16	22213	1613	1613	0.999	0.054	100
7.21-42.47	12.291	0.043	38.11	11492	939	935	0.999	0.044	99.6

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Phasing

Phasing

Method:

molecular replacement

Phasing MR

	Highest resolution	Lowest resolution
Rotation	3.58 Å	44.52 Å
Translation	3.58 Å	44.52 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: 1.61→42.47 Å / SU ML: 0.18 / Cross valid method: THROUGHOUT / σ(F): 1.36 / Phase error: 19.35 / Stereochemistry target values: ML

	R_factor	Num. reflection	% reflection
R_free	0.1857	3698	5 %
R_work	0.1509	70230	-
obs	0.1526	73928	99.57 %

Solvent computation

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

Displacement parameters

B_iso max: 64.09 Å² / B_iso mean: 24.5907 Å² / B_iso min: 12.51 Å²

Refinement step

Cycle: final / Resolution: 1.61→42.47 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4748	0	147	838	5733
B_iso mean	-	-	30.49	35.01	-
Num. residues	-	-	-	-	610

LS refinement shell

Refine-ID: X-RAY DIFFRACTION / R_factor R_free error: 0 / Total num. of bins used: 26

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Num. reflection all	% reflection obs (%)
1.61-1.63	0.3282	127	0.2843	2401	2528	90
1.63-1.66	0.2725	141	0.2383	2675	2816	100
1.66-1.68	0.2405	141	0.216	2689	2830	100
1.68-1.7	0.2508	141	0.2379	2678	2819	100
1.7-1.73	0.2645	141	0.2351	2679	2820	100
1.73-1.76	0.2818	142	0.2099	2690	2832	100
1.76-1.79	0.221	141	0.2019	2672	2813	100
1.79-1.82	0.2338	141	0.1859	2692	2833	100
1.82-1.86	0.2212	142	0.1706	2677	2819	100
1.86-1.89	0.2025	142	0.1715	2707	2849	100
1.89-1.94	0.2278	141	0.1824	2673	2814	100
1.94-1.98	0.2411	142	0.1616	2705	2847	100
1.98-2.03	0.1918	142	0.1515	2700	2842	100
2.03-2.09	0.1862	142	0.1519	2681	2823	100
2.09-2.15	0.1948	141	0.1533	2687	2828	100
2.15-2.22	0.1815	143	0.1462	2710	2853	100
2.22-2.29	0.2008	142	0.1542	2701	2843	100
2.3-2.39	0.2089	142	0.1434	2686	2828	100
2.39-2.5	0.1803	143	0.1495	2726	2869	100
2.5-2.63	0.1678	143	0.1482	2711	2854	100
2.63-2.79	0.1899	143	0.144	2735	2878	100
2.79-3.01	0.1878	144	0.1489	2731	2875	100
3.01-3.31	0.1796	144	0.1473	2737	2881	100
3.31-3.79	0.1685	146	0.1336	2779	2925	100
3.79-4.77	0.1308	147	0.1141	2786	2933	100
4.77-42.47	0.1653	154	0.14	2922	3076	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	1.3498	0.8125	-0.2057	1.538	-0.028	1.7675	0.023	-0.1615	-0.1543	-0.0745	0.0267	-0.035	0.268	0.0586	-0.0394	0.187	0.0125	-0.0469	0.1488	-0.0004	0.2288	-4.7255	16.34	-9.8134
2	2.1547	-0.0432	-0.6275	1.3401	0.2568	1.938	-0.0104	-0.1898	-0.2848	0.1594	-0.0159	-0.0746	0.4369	-0.0153	0.0388	0.2749	-0.0238	-0.0493	0.173	0.0462	0.2674	-5.9911	10.1924	-0.7904
3	0.8591	0.3031	0.1554	0.5857	0.1131	0.8844	0.0309	-0.0892	0.0093	0.0082	0.0045	0.0625	0.0145	-0.0907	-0.0359	0.1317	0.0072	0.0002	0.1593	0.0036	0.1843	-9.055	27.9208	-6.2108
4	0.5358	0.0161	0.1324	0.8657	0.3043	0.836	-0.0295	0.1014	-0.0485	-0.0064	0.1022	-0.0421	0.0184	0.0839	-0.0484	0.1493	-0.0039	-0.002	0.1565	-0.0017	0.1818	-6.0633	22.6927	-20.4548
5	0.9851	0.5375	0.347	2.1389	0.6977	1.0469	-0.0097	0.2279	-0.0175	-0.1985	0.0362	-0.0496	-0.0088	0.192	-0.0408	0.2261	-0.0087	0.0091	0.2183	-0.0203	0.1902	-1.6881	26.6757	-28.9556
6	0.5064	-0.0238	0.0648	1.7764	-0.063	0.6487	0.0427	0.2035	-0.0772	-0.0317	-0.0214	0.0443	-0.0579	0.0273	-0.0631	0.1721	0.0001	0.0052	0.1762	-0.0159	0.1956	-1.6408	21.8827	-20.1211
7	1.4199	0.701	0.2035	2.14	0.2156	1.1113	0.0474	0.0241	-0.1768	-0.1549	-0.0403	-0.0004	0.1579	0.2504	-0.006	0.2102	0.0421	-0.005	0.2508	-0.0427	0.2248	2.6203	14.7884	-27.3356
8	0.453	0.2098	-0.0379	0.8713	0.0991	1.1079	0.0219	-0.0308	-0.2271	0.0678	-0.0021	-0.0684	0.2843	0.0921	-0.0079	0.2146	0.0344	-0.0234	0.1698	-0.023	0.2708	-1.324	9.6887	-15.0194
9	0.6503	-0.0181	-0.0229	1.4322	-0.0234	2.0908	-0.0287	-0.0148	-0.16	-0.0556	0.027	0.1248	0.3548	-0.3428	-0.0494	0.2194	-0.0089	0.0058	0.2504	0.0211	0.1983	-38.0834	19.0837	-34.0843
10	2.3271	-0.1784	-0.5224	1.4352	-0.2821	2.3225	0.0547	0.223	-0.1786	-0.2264	-0.0329	0.0738	0.6144	-0.4878	0.0107	0.3257	-0.0636	-0.0133	0.245	-0.0019	0.1729	-37.688	14.4557	-41.2377
11	0.4796	-0.1333	0.0452	0.6527	-0.1366	1.5459	-0.047	0.0045	-0.0386	-0.0625	0.0438	-0	0.2069	-0.0319	-0.0013	0.1754	0.0015	-0.0002	0.1958	-0.0048	0.1819	-29.5981	24.2947	-44.2858
12	0.3282	-0.0104	-0.1853	0.6297	-0.2178	1.2306	-0.0336	-0.0211	0.046	0.0984	0.0652	0.027	-0.1546	-0.122	-0.0295	0.1749	0.0202	0.0013	0.1735	0.0069	0.1881	-31.3978	33.7164	-34.0244
13	0.3586	0.4124	-0.2709	1.076	-0.1317	1.0249	0.004	-0.0385	-0.0245	0.2552	0.0522	-0.0684	0.0568	-0.106	-0.0456	0.2428	0.0257	0.0355	0.2148	0.0241	0.192	-33.7215	22.4907	-16.0067
14	0.7796	0.0905	-0.0756	2.325	-0.1038	0.9927	0.0228	0.021	-0.0375	0.2336	-0.0687	0.1088	0.0165	-0.3458	0.0554	0.2679	0.0216	0.0281	0.2406	0.0321	0.1979	-37.8025	24.1901	-23.8741
15	2.1601	-0.7478	0.1546	2.2313	-0.6753	2.0409	0.014	-0.0616	-0.0479	0.1482	0.0766	0.1179	0.2622	-0.5245	-0.0693	0.2321	-0.0299	0.0348	0.2921	0.0366	0.2159	-43.0436	19.1125	-16.1258
16	0.8197	-0.1052	-0.2605	1.2128	-0.126	2.2222	0.0124	0.1196	-0.2095	-0.1656	-0.0064	0.1205	0.6886	-0.5905	-0.032	0.3721	-0.1042	0.0083	0.2762	0.0098	0.2318	-40.9386	12.5662	-28.6849

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 43 )	A	21 - 43
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 44 through 185 )	A	44 - 185
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 186 through 216 )	A	186 - 216
5	X-RAY DIFFRACTION	5	chain 'A' and (resid 217 through 231 )	A	217 - 231
6	X-RAY DIFFRACTION	6	chain 'A' and (resid 232 through 246 )	A	232 - 246
7	X-RAY DIFFRACTION	7	chain 'A' and (resid 247 through 263 )	A	247 - 263
8	X-RAY DIFFRACTION	8	chain 'A' and (resid 264 through 305 )	A	264 - 305
9	X-RAY DIFFRACTION	9	chain 'B' and (resid 1 through 20 )	B	1 - 20
10	X-RAY DIFFRACTION	10	chain 'B' and (resid 21 through 52 )	B	21 - 52
11	X-RAY DIFFRACTION	11	chain 'B' and (resid 53 through 105 )	B	53 - 105
12	X-RAY DIFFRACTION	12	chain 'B' and (resid 106 through 200 )	B	106 - 200
13	X-RAY DIFFRACTION	13	chain 'B' and (resid 201 through 231 )	B	201 - 231
14	X-RAY DIFFRACTION	14	chain 'B' and (resid 232 through 246 )	B	232 - 246
15	X-RAY DIFFRACTION	15	chain 'B' and (resid 247 through 263 )	B	247 - 263
16	X-RAY DIFFRACTION	16	chain 'B' and (resid 264 through 305 )	B	264 - 305

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