PDB-8j51: Crystal structure of Flavihumibacter petaseus GH31 alpha-galactos...

ID or keywords:

Entry

Database: PDB / ID: 8j51

Title

Crystal structure of Flavihumibacter petaseus GH31 alpha-galactosidase in complex with galactose

Components

GH31 alpha-galactosidase

Keywords

HYDROLASE / Glycoside hydrolase / (beta/alpha)8 barrel / Globo sphingolipid / glycolipid / Carbohydrate

Function / homology

Function and homology information

hydrolase activity, hydrolyzing O-glycosyl compounds / carbohydrate metabolic process
Similarity search - Function

Biological species

Flavihumibacter petaseus NBRC 106054 (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.15 Å

Authors

Ikegaya, M. / Miyazaki, T.

Funding support

Japan, 1items

Citation

Journal: Febs J. / Year: 2023
Title: Structure-function analysis of bacterial GH31 alpha-galactosidases specific for alpha-(1→4)-galactobiose.
Authors: Ikegaya, M. / Park, E.Y. / Miyazaki, T.

History

Deposition	Apr 21, 2023	Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0	Jul 26, 2023	Provider: repository / Type: Initial release
Revision 1.1	Nov 1, 2023	Group: Data collection / Database references Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / citation / citation_author Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	8j51.cif.gz	1 MB	Display	PDBx/mmCIF format
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Validation report

Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/j5/8j51 ftp://data.pdbj.org/pub/pdb/validation_reports/j5/8j51	HTTPS FTP

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Related structure data

Related structure data	8j50C 8j52C 8j53C 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
Related items in Molecule of the Month	#156 - Dec 2012 ABO Blood Type Glycosyltransferases similarity (1) #198 - Jun 2016 Beta-galactosidase similarity (1)

Deposited unit

A: GH31 alpha-galactosidase

B: GH31 alpha-galactosidase

hetero molecules

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

1

Idetical with deposited unit
defined by author
Evidence: gel filtration

Unit cell

Length a, b, c (Å)	71.357, 72.371, 80.884
Angle α, β, γ (deg.)	101.822, 105.114, 103.668
Int Tables number	1
Space group name H-M	P1

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Protein , 1 types, 2 molecules AB

#1: Protein	GH31 alpha-galactosidase Mass: 60973.340 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Flavihumibacter petaseus NBRC 106054 (bacteria) Strain: NBRC 106054 / Gene: FPE01S_01_01430 / Plasmid: pET28a / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / References: UniProt: A0A0E9MUN5, alpha-galactosidase

#1: Protein

GH31 alpha-galactosidase

Mass: 60973.340 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.)

Flavihumibacter petaseus NBRC 106054 (bacteria)
Strain: NBRC 106054 / Gene: FPE01S_01_01430 / Plasmid: pET28a / Production host:

Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / References: UniProt: A0A0E9MUN5, alpha-galactosidase

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Sugars , 2 types, 4 molecules

#2: Sugar

ChemComp-GLA / alpha-D-galactopyranose / alpha-D-galactose / D-galactose / galactose / ALPHA D-GALACTOSE

Type: D-saccharide, alpha linking / Mass: 180.156 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₆H₁₂O₆ / Feature type: SUBJECT OF INVESTIGATION

#3: Sugar

ChemComp-GAL / beta-D-galactopyranose / beta-D-galactose / D-galactose / galactose

Type: D-saccharide, beta linking / Mass: 180.156 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₆H₁₂O₆ / Feature type: SUBJECT OF INVESTIGATION

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Non-polymers , 3 types, 400 molecules

#4: Chemical	ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL Mass: 62.068 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C₂H₆O₂
#5: Chemical	ChemComp-CL / CHLORIDE ION Mass: 35.453 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Cl
#6: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 386 / Source method: isolated from a natural source / Formula: H₂O

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Details

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: 3.08 Å³/Da / Density % sol: 60.07 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 6 / Details: 1 M sodium chloride, 50 mM MES-NaOH, pH 6.0

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: Photon Factory / Beamline: BL-5A / Wavelength: 1 Å
Detector	Type: DECTRIS PILATUS3 S 6M / Detector: PIXEL / Date: Jun 25, 2022
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 1 Å / Relative weight: 1
Reflection	Resolution: 2.15→46.01 Å / Num. obs: 77597 / % possible obs: 98 % / Redundancy: 3.5 % / CC_1/2: 0.993 / R_merge(I) obs: 0.097 / R_rim(I) all: 0.138 / Net I/σ(I): 9.9
Reflection shell	Resolution: 2.15→2.19 Å / Redundancy: 3.5 % / R_merge(I) obs: 0.487 / Mean I/σ(I) obs: 2.1 / Num. unique obs: 4574 / CC_1/2: 0.739 / R_pim(I) all: 0.487 / R_rim(I) all: 0.689 / % possible all: 97.1

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0405	refinement
XDS		data reduction
Aimless		data scaling
MOLREP		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 2.15→43.695 Å / Cor.coef. F_o:F_c: 0.963 / Cor.coef. F_o:F_c free: 0.923 / WR_factor R_free: 0.198 / WR_factor R_work: 0.139 / SU B: 10.904 / SU ML: 0.123 / Average fsc free: 0.9675 / Average fsc work: 0.9871 / Cross valid method: FREE R-VALUE / ESU R Free: 0.167
Details: Hydrogens have been added in their riding positions

	R_factor	Num. reflection	% reflection
R_free	0.2224	3948	5.088 %
R_work	0.152	73645	-
all	0.155	-	-
obs	-	77593	97.972 %

Solvent computation

Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: BABINET MODEL PLUS MASK

Displacement parameters

B_iso mean: 34.168 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	0.842 Å²	0.745 Å²	0.738 Å²
2-	-	-0.828 Å²	0.221 Å²
3-	-	-	-0.705 Å²

Refinement step

Cycle: LAST / Resolution: 2.15→43.695 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	8114	0	98	386	8598

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.008	0.012	8431
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.016	7888
X-RAY DIFFRACTION	r_angle_refined_deg	1.439	1.643	11410
X-RAY DIFFRACTION	r_angle_other_deg	0.496	1.568	18129
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	7.099	5	1020
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	9.488	5	60
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	15.267	10	1371
X-RAY DIFFRACTION	r_dihedral_angle_6_deg	16.395	10	397
X-RAY DIFFRACTION	r_chiral_restr	0.067	0.2	1196
X-RAY DIFFRACTION	r_gen_planes_refined	0.007	0.02	9900
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	2072
X-RAY DIFFRACTION	r_nbd_refined	0.208	0.2	1621
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.2	0.2	7504
X-RAY DIFFRACTION	r_nbtor_refined	0.186	0.2	4039
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.082	0.2	4355
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.137	0.2	365
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_other	0.051	0.2	3
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.253	0.2	8
X-RAY DIFFRACTION	r_nbd_other	0.23	0.2	66
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.108	0.2	12
X-RAY DIFFRACTION	r_mcbond_it	2.057	3.412	4080
X-RAY DIFFRACTION	r_mcbond_other	2.056	3.412	4080
X-RAY DIFFRACTION	r_mcangle_it	2.957	6.123	5094
X-RAY DIFFRACTION	r_mcangle_other	2.957	6.125	5095
X-RAY DIFFRACTION	r_scbond_it	2.292	3.679	4351
X-RAY DIFFRACTION	r_scbond_other	2.291	3.679	4352
X-RAY DIFFRACTION	r_scangle_it	3.29	6.644	6314
X-RAY DIFFRACTION	r_scangle_other	3.29	6.644	6315
X-RAY DIFFRACTION	r_lrange_it	4.15	33.569	9492
X-RAY DIFFRACTION	r_lrange_other	4.11	33.565	9461
X-RAY DIFFRACTION	r_rigid_bond_restr	3.81	3	16319

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
2.15-2.206	0.273	280	0.192	5454	0.196	5914	0.953	0.978	96.9564	0.18
2.206-2.266	0.276	329	0.181	5175	0.186	5663	0.951	0.981	97.1923	0.168
2.266-2.332	0.26	304	0.167	5082	0.172	5552	0.957	0.984	97.0101	0.15
2.332-2.403	0.244	270	0.153	4951	0.158	5360	0.963	0.986	97.4067	0.136
2.403-2.482	0.219	272	0.139	4806	0.143	5205	0.972	0.989	97.56	0.122
2.482-2.568	0.226	265	0.14	4680	0.144	5064	0.969	0.989	97.6501	0.122
2.568-2.665	0.214	238	0.155	4501	0.158	4854	0.97	0.987	97.6308	0.133
2.665-2.773	0.255	203	0.151	4420	0.156	4722	0.966	0.989	97.9034	0.129
2.773-2.896	0.246	191	0.155	4215	0.159	4490	0.965	0.988	98.1292	0.132
2.896-3.037	0.244	202	0.14	3985	0.145	4263	0.965	0.989	98.2172	0.117
3.037-3.2	0.241	222	0.146	3815	0.151	4111	0.965	0.988	98.2	0.124
3.2-3.393	0.229	181	0.154	3619	0.158	3865	0.972	0.987	98.3182	0.135
3.393-3.626	0.212	166	0.15	3412	0.153	3629	0.975	0.989	98.5947	0.138
3.626-3.914	0.183	171	0.141	3167	0.143	3380	0.981	0.991	98.7574	0.13
3.914-4.284	0.182	146	0.129	2955	0.131	3134	0.984	0.992	98.947	0.119
4.284-4.784	0.187	143	0.133	2634	0.136	2801	0.982	0.992	99.1432	0.123
4.784-5.514	0.191	107	0.151	2374	0.153	2501	0.985	0.993	99.2003	0.141
5.514-6.726	0.217	90	0.176	2007	0.178	2110	0.974	0.988	99.3839	0.159
6.726-9.401	0.199	92	0.152	1524	0.155	1626	0.977	0.987	99.385	0.143
9.401-43.695	0.238	76	0.226	866	0.227	946	0.983	0.983	99.5772	0.244

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