PDB-8j5m: Structure of GH1 Br2 beta-glucosidase E350G mutant from bovine ru...

ID or keywords:

Entry

Database: PDB / ID: 8j5m

Title

Structure of GH1 Br2 beta-glucosidase E350G mutant from bovine rumen metagenome

Components

Beta-glucosidase

Keywords

HYDROLASE /

Glycoside hydrolase / GH1 /

metagenome /

beta-glucosidase

Function / homology

Function and homology information

scopolin beta-glucosidase activity /

beta-glucosidase activity /

beta-glucosidase / cellulose catabolic process
Similarity search - Function

Biological species

uncultured bacterium (environmental samples)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.621 Å

Authors

Kaenying, W. / Kongsaeree, P.T. / Tagami, T.

Funding support

Thailand, 1items

Citation

Journal: Heliyon / Year: 2023
Title: Structural and mutational analysis of glycoside hydrolase family 1 Br2 beta-glucosidase derived from bovine rumen metagenome.
Authors: Kaenying, W. / Tagami, T. / Suwan, E. / Pitsanuwong, C. / Chomngam, S. / Okuyama, M. / Kongsaeree, P. / Kimura, A. / Kongsaeree, P.T.

History

Deposition	Apr 23, 2023	Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0	Nov 22, 2023	Provider: repository / Type: Initial release
Revision 1.1	Dec 13, 2023	Group: Database references / Category: citation / citation_author Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name
Revision 1.2	Jan 3, 2024	Group: Database references / Category: citation Item: _citation.page_first / _citation.page_last / _citation.title

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	8j5m.cif.gz	429.4 KB	Display	PDBx/mmCIF format
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PDBx/mmJSON format	8j5m.json.gz		Tree view	PDBx/mmJSON format
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Validation report

Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/j5/8j5m ftp://data.pdbj.org/pub/pdb/validation_reports/j5/8j5m	HTTPS FTP

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Related structure data

Related structure data	8j3mC 8j5lC C: citing same article (ref.)
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: Beta-glucosidase

B: Beta-glucosidase

D: Beta-glucosidase

C: Beta-glucosidase

hetero molecules

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

1

Idetical with deposited unit
defined by author&software

Unit cell

Length a, b, c (Å)	105.297, 113.605, 180.860
Angle α, β, γ (deg.)	90.000, 90.000, 90.000
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
3	2	A
4	2	C
5	3	A
6	3	D
7	4	B
8	4	C
9	5	B
10	5	D
11	6	C
12	6	D

NCS domain segments:

End auth comp-ID: LEU / End label comp-ID: LEU / Refine code: 1

Dom-ID	Component-ID	Ens-ID	Beg auth comp-ID	Beg label comp-ID	Auth asym-ID	Label asym-ID	Auth seq-ID	Label seq-ID
1	1	1	LEU	LEU	A	A	-6 - 445	14 - 465
2	1	1	LEU	LEU	B	B	-6 - 445	14 - 465
3	2	2	GLY	GLY	A	A	2 - 444	22 - 464
4	2	2	GLY	GLY	D	C	2 - 444	22 - 464
5	3	3	GLY	GLY	A	A	2 - 444	22 - 464
6	3	3	GLY	GLY	C	D	2 - 444	22 - 464
7	4	4	GLY	GLY	B	B	2 - 444	22 - 464
8	4	4	GLY	GLY	D	C	2 - 444	22 - 464
9	5	5	GLY	GLY	B	B	2 - 444	22 - 464
10	5	5	GLY	GLY	C	D	2 - 444	22 - 464
11	6	6	GLY	GLY	D	C	2 - 445	22 - 465
12	6	6	GLY	GLY	C	D	2 - 445	22 - 465

NCS ensembles :

ID	Details
1	Local NCS retraints between domains: 1 2
2	Local NCS retraints between domains: 3 4
3	Local NCS retraints between domains: 5 6
4	Local NCS retraints between domains: 7 8
5	Local NCS retraints between domains: 9 10
6	Local NCS retraints between domains: 11 12

#1: Protein	Beta-glucosidase / Mass: 53016.680 Da / Num. of mol.: 4 / Mutation: E350G Source method: isolated from a genetically manipulated source Details: bovine rumen metagenome Source: (gene. exp.) uncultured bacterium (environmental samples) Gene: BG / Plasmid: pET15b / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0A1S5SJM8
#2: Chemical	ChemComp-ACT / ACETATE ION / Acetate Mass: 59.044 Da / Num. of mol.: 9 / Source method: obtained synthetically / Formula: C₂H₃O₂
#3: Chemical	... ChemComp-SO4 / SULFATE ION / Sulfate Mass: 96.063 Da / Num. of mol.: 33 / Source method: obtained synthetically / Formula: SO₄
#4: Water	ChemComp-HOH / water / Water Mass: 18.015 Da / Num. of mol.: 2107 / 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: 2.58 Å³/Da / Density % sol: 51.77 %
Crystal grow	Temperature: 277 K / Method: vapor diffusion, hanging drop / pH: 7.4 Details: 0.1M sodium acetate trihydrate pH 7.4, 1.6M ammonium sulfate

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: NSRRC / Beamline: BL13C1 / Wavelength: 0.975 Å
Detector	Type: ADSC QUANTUM 315r / Detector: CCD / Date: Jun 6, 2018
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 0.975 Å / Relative weight: 1
Reflection	Resolution: 1.62→50 Å / Num. obs: 271392 / % possible obs: 99.2 % / Redundancy: 4.9 % / B_iso Wilson estimate: 29.082 Å² / CC_1/2: 0.999 / R_rim(I) all: 0.074 / R_sym value: 0.066 / Net I/σ(I): 13.18
Reflection shell	Resolution: 1.62→1.72 Å / Redundancy: 4.8 % / Mean I/σ(I) obs: 1.96 / Num. unique obs: 42595 / CC_1/2: 0.745 / R_rim(I) all: 0.816 / R_sym value: 0.727 / % possible all: 97.1

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0267	refinement
XDS		data reduction
XDS		data scaling
PHENIX		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 1.621→42.237 Å / Cor.coef. F_o:F_c: 0.974 / Cor.coef. F_o:F_c free: 0.963 / SU B: 1.795 / SU ML: 0.059 / Cross valid method: FREE R-VALUE / ESU R: 0.076 / ESU R Free: 0.076
Details: Hydrogens have been added in their riding positions

	R_factor	Num. reflection	% reflection
R_free	0.1877	2001	0.7 %
R_work	0.1609	269390	-
all	0.161	-	-
obs	-	269390	99.17 %

Solvent computation

Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK BULK SOLVENT

Displacement parameters

B_iso mean: 23.012 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	-0.56 Å²	0 Å²	0 Å²
2-	-	0.802 Å²	0 Å²
3-	-	-	-0.242 Å²

Refinement step

Cycle: LAST / Resolution: 1.621→42.237 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	14459	0	201	2107	16767

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.008	0.013	15097
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.017	13588
X-RAY DIFFRACTION	r_angle_refined_deg	1.381	1.635	20498
X-RAY DIFFRACTION	r_angle_other_deg	1.435	1.58	31294
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	6.656	5	1800
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	33.514	23.023	817
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	11.59	15	2404
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	17.441	15	66
X-RAY DIFFRACTION	r_chiral_restr	0.076	0.2	1836
X-RAY DIFFRACTION	r_gen_planes_refined	0.007	0.02	17256
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	3684
X-RAY DIFFRACTION	r_nbd_refined	0.213	0.2	3128
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.173	0.2	13364
X-RAY DIFFRACTION	r_nbtor_refined	0.177	0.2	7523
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.075	0.2	6549
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.139	0.2	1835
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_other	0.105	0.2	2
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.206	0.2	32
X-RAY DIFFRACTION	r_nbd_other	0.204	0.2	79
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.153	0.2	40
X-RAY DIFFRACTION	r_mcbond_it	1.46	2.172	7203
X-RAY DIFFRACTION	r_mcbond_other	1.458	2.17	7196
X-RAY DIFFRACTION	r_mcangle_it	2.049	3.253	8996
X-RAY DIFFRACTION	r_mcangle_other	2.049	3.253	8997
X-RAY DIFFRACTION	r_scbond_it	2.426	2.47	7894
X-RAY DIFFRACTION	r_scbond_other	2.426	2.47	7895
X-RAY DIFFRACTION	r_scangle_it	3.607	3.606	11502
X-RAY DIFFRACTION	r_scangle_other	3.607	3.606	11503
X-RAY DIFFRACTION	r_lrange_it	5.382	27.326	18713
X-RAY DIFFRACTION	r_lrange_other	5.381	27.329	18714
X-RAY DIFFRACTION	r_ncsr_local_group_1	0.053	0.05	15925
X-RAY DIFFRACTION	r_ncsr_local_group_2	0.07	0.05	15465
X-RAY DIFFRACTION	r_ncsr_local_group_3	0.065	0.05	15491
X-RAY DIFFRACTION	r_ncsr_local_group_4	0.074	0.05	15470
X-RAY DIFFRACTION	r_ncsr_local_group_5	0.069	0.05	15502
X-RAY DIFFRACTION	r_ncsr_local_group_6	0.072	0.05	15573

Refine LS restraints NCS

Ens-ID	Dom-ID	Auth asym-ID	Refine-ID	Type	Rms dev position (Å)	Weight position
1	1	A	X-RAY DIFFRACTION	Local ncs	0.05274	0.05009
1	2	B	X-RAY DIFFRACTION	Local ncs	0.05274	0.05009
2	3	A	X-RAY DIFFRACTION	Local ncs	0.06994	0.05009
2	4	C	X-RAY DIFFRACTION	Local ncs	0.06994	0.05009
3	5	A	X-RAY DIFFRACTION	Local ncs	0.06477	0.05009
3	6	D	X-RAY DIFFRACTION	Local ncs	0.06477	0.05009
4	7	B	X-RAY DIFFRACTION	Local ncs	0.07376	0.05009
4	8	C	X-RAY DIFFRACTION	Local ncs	0.07376	0.05009
5	9	B	X-RAY DIFFRACTION	Local ncs	0.06924	0.05009
5	10	D	X-RAY DIFFRACTION	Local ncs	0.06924	0.05009
6	11	C	X-RAY DIFFRACTION	Local ncs	0.07155	0.05009
6	12	D	X-RAY DIFFRACTION	Local ncs	0.07155	0.05009

LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Total num. of bins used: 20

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	R_factor all	Num. reflection all	Fsc free	Fsc work	% reflection obs (%)	WR_factor R_work
1.621-1.663	0.282	138	0.268	18626	0.268	20039	0.833	0.824	93.6374	0.248
1.663-1.708	0.262	140	0.246	19423	0.246	19564	0.848	0.862	99.9949	0.221
1.708-1.758	0.239	153	0.224	18890	0.224	19048	0.884	0.892	99.9737	0.195
1.758-1.812	0.237	121	0.211	18319	0.211	18442	0.894	0.91	99.9892	0.181
1.812-1.871	0.239	136	0.202	17776	0.202	17913	0.9	0.918	99.9944	0.171
1.871-1.937	0.215	134	0.187	17221	0.188	17377	0.933	0.939	99.8734	0.159
1.937-2.01	0.194	120	0.18	16609	0.181	16741	0.947	0.948	99.9283	0.153
2.01-2.092	0.196	109	0.17	15995	0.17	16137	0.952	0.956	99.7955	0.147
2.092-2.184	0.183	125	0.165	15380	0.165	15517	0.956	0.959	99.9227	0.144
2.184-2.291	0.176	111	0.153	14663	0.153	14812	0.961	0.965	99.7434	0.134
2.291-2.414	0.156	98	0.147	13996	0.147	14120	0.962	0.967	99.8159	0.131
2.414-2.56	0.174	105	0.144	13226	0.145	13356	0.965	0.969	99.8128	0.131
2.56-2.736	0.215	89	0.153	12455	0.154	12597	0.948	0.964	99.5793	0.143
2.736-2.954	0.162	87	0.157	11618	0.157	11754	0.962	0.963	99.5831	0.151
2.954-3.235	0.172	75	0.159	10719	0.159	10863	0.966	0.966	99.3648	0.159
3.235-3.614	0.205	77	0.152	9680	0.152	9862	0.947	0.965	98.9353	0.161
3.614-4.168	0.157	60	0.133	8547	0.133	8726	0.974	0.975	98.6363	0.15
4.168-5.092	0.156	56	0.124	7283	0.124	7452	0.982	0.983	98.4836	0.144
5.092-7.149	0.162	40	0.15	5696	0.151	5860	0.969	0.975	97.884	0.173
7.149-42.237	0.235	27	0.163	3268	0.163	3455	0.945	0.968	95.369	0.204

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