PDB-8agn: Cyclohexane epoxide low pH soak of epoxide hydrolase from metagen...

ID or keywords:

Entry

Database: PDB / ID: 8agn

Title

Cyclohexane epoxide low pH soak of epoxide hydrolase from metagenomic source ch65

Components

Alpha/beta epoxide hydrolase

Keywords

HYDROLASE / Alpha/beta hydrolase fold / complex with substrate / limonene epoxide

Function / homology

Epoxide hydrolase-like / alpha/beta hydrolase fold / Alpha/beta hydrolase fold-1 / Alpha/Beta hydrolase fold / hydrolase activity / (1R,6S)-7-oxabicyclo[4.1.0]heptane / DI(HYDROXYETHYL)ETHER / Alpha/beta epoxide hydrolase

Function and homology information

Biological species

metagenome (others)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.957 Å

Authors

Isupov, M.N. / De Rose, S.A. / Mitchell, D. / Littlechild, J.A.

Funding support

European Union, 1items

Citation

Journal: To Be Published
Title: Complexes of epoxide hydrolase from metagenomic source ch65
Authors: Isupov, M.N. / De Rose, S.A. / Mitchell, D. / Littlechild, J.A. / Parker, E. / Ferrandi, E. / Guazzelli, E. / Monti, D.

History

Deposition	Jul 20, 2022	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Aug 16, 2023	Provider: repository / Type: Initial release

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	8agn.cif.gz	268.5 KB	Display	PDBx/mmCIF format
PDB format	pdb8agn.ent.gz		Display	PDB format
PDBx/mmJSON format	8agn.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	8agn_validation.pdf.gz	2.5 MB	Display	wwPDB validaton report
Full document	8agn_full_validation.pdf.gz	2.5 MB	Display
Data in XML	8agn_validation.xml.gz	46.4 KB	Display
Data in CIF	8agn_validation.cif.gz	64.2 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/ag/8agn ftp://data.pdbj.org/pub/pdb/validation_reports/ag/8agn	HTTPS FTP

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Related structure data

Related structure data	8agmC 8agpC 8agsC 5ng7S C: citing same article (ref.) 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	#165 - Sep 2013 Designed Protein Cages similarity (3) #220 - Apr 2018 Dehalogenases similarity (3) #216 - Dec 2017 Biodegradable Plastic similarity (1)

Deposited unit

AAA: Alpha/beta epoxide hydrolase

BBB: Alpha/beta epoxide hydrolase

CCC: Alpha/beta epoxide hydrolase

DDD: Alpha/beta epoxide hydrolase

hetero molecules

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

1

AAA: Alpha/beta epoxide hydrolase

BBB: Alpha/beta epoxide hydrolase

hetero molecules

defined by author
Evidence: gel filtration
70.9 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

CCC: Alpha/beta epoxide hydrolase

DDD: Alpha/beta epoxide hydrolase

hetero molecules

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

Unit cell

Length a, b, c (Å)	95.851, 47.330, 142.611
Angle α, β, γ (deg.)	90.000, 91.707, 90.000
Int Tables number	4
Space group name H-M	P12₁1

Noncrystallographic symmetry (NCS)

NCS domain:

ID	Ens-ID	Details
1	1	AAA
2	1	BBB
3	2	AAA
4	2	CCC
5	3	AAA
6	3	DDD
7	4	BBB
8	4	CCC
9	5	BBB
10	5	DDD
11	6	CCC
12	6	DDD

NCS domain segments:

Dom-ID	Component-ID	Ens-ID	Beg auth comp-ID	Beg label comp-ID	End auth comp-ID	End label comp-ID	Auth asym-ID	Label asym-ID	Auth seq-ID	Label seq-ID
1	1	1	MET	MET	LEU	LEU	AAA	A	4 - 289	4 - 289
2	2	1	MET	MET	LEU	LEU	BBB	B	4 - 289	4 - 289
3	3	2	MET	MET	LEU	LEU	AAA	A	4 - 289	4 - 289
4	4	2	MET	MET	LEU	LEU	CCC	C	4 - 289	4 - 289
5	5	3	MET	MET	LEU	LEU	AAA	A	4 - 289	4 - 289
6	6	3	MET	MET	LEU	LEU	DDD	D	4 - 289	4 - 289
7	7	4	GLU	GLU	LEU	LEU	BBB	B	3 - 289	3 - 289
8	8	4	GLU	GLU	LEU	LEU	CCC	C	3 - 289	3 - 289
9	9	5	GLU	GLU	LEU	LEU	BBB	B	3 - 289	3 - 289
10	10	5	GLU	GLU	LEU	LEU	DDD	D	3 - 289	3 - 289
11	11	6	GLU	GLU	SER	SER	CCC	C	3 - 291	3 - 291
12	12	6	GLU	GLU	SER	SER	DDD	D	3 - 291	3 - 291

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	Alpha/beta epoxide hydrolase Mass: 34940.449 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) metagenome (others) Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: A0A1U9WZ52
#2: Chemical	ChemComp-7EX / (1R,6S)-7-oxabicyclo[4.1.0]heptane Mass: 98.143 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: C₆H₁₀O / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical	ChemComp-PEG / DI(HYDROXYETHYL)ETHER Mass: 106.120 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C₄H₁₀O₃
#4: Chemical	ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL Mass: 62.068 Da / Num. of mol.: 11 / Source method: obtained synthetically / Formula: C₂H₆O₂
#5: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 272 / 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: 2.31 Å³/Da / Density % sol: 46.83 %
Crystal grow	Temperature: 291 K / Method: microbatch / pH: 7.5 Details: Morpheus crystallisation kit condition H8, 25% v/v MPD; 25% PEG 1000; 25% w/v PEG 3350 . The crystal was soaked for 2 hours in cryoprotectant containing 1mM of cyclohexane oxide at pH 4.5.

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: Diamond / Beamline: I04 / Wavelength: 0.9795 Å
Detector	Type: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Oct 20, 2018
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 0.9795 Å / Relative weight: 1
Reflection	Resolution: 1.957→142.548 Å / Num. obs: 102758 / % possible obs: 99.7 % / Redundancy: 3.4 % / CC_1/2: 0.999 / Net I/σ(I): 10
Reflection shell	Resolution: 1.957→1.991 Å / Redundancy: 3.4 % / Num. unique obs: 4581 / CC_1/2: 0.303 / % possible all: 99.7

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0267	refinement
XDS		data reduction
autoPROC		data processing
XSCALE		data scaling
MOLREP		phasing
Coot		model building
PARROT		phasing
BUSTER		refinement

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 5ng7

5ng7

Resolution: 1.957→142.548 Å / Cor.coef. F_o:F_c: 0.97 / Cor.coef. F_o:F_c free: 0.95 / SU B: 9.229 / SU ML: 0.224 / Cross valid method: FREE R-VALUE / ESU R: 0.19 / ESU R Free: 0.175 / Details: Hydrogens have not been used

	R_factor	Num. reflection	% reflection
R_free	0.2587	4502	4.849 %
R_work	0.2055	88351	-
all	0.208	-	-
obs	-	92853	99.681 %

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: 54.15 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	-3.405 Å²	-0 Å²	-0.919 Å²
2-	-	5.268 Å²	-0 Å²
3-	-	-	-1.805 Å²

Refinement step

Cycle: LAST / Resolution: 1.957→142.548 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	9543	0	135	272	9950

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.01	0.012	10258
X-RAY DIFFRACTION	r_angle_refined_deg	1.64	1.64	13933
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	6.608	5	1245
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	33.967	22.156	538
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	17.455	15	1850
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	21.336	15	60
X-RAY DIFFRACTION	r_chiral_restr	0.104	0.2	1292
X-RAY DIFFRACTION	r_gen_planes_refined	0.009	0.02	7652
X-RAY DIFFRACTION	r_nbd_refined	0.214	0.2	4627
X-RAY DIFFRACTION	r_nbtor_refined	0.304	0.2	6785
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.139	0.2	399
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.222	0.2	58
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.106	0.2	16
X-RAY DIFFRACTION	r_mcbond_it	8.851	13.63	4723
X-RAY DIFFRACTION	r_mcangle_it	10.434	22.891	5921
X-RAY DIFFRACTION	r_scbond_it	10.857	14.897	5535
X-RAY DIFFRACTION	r_scangle_it	12.855	24.216	7943
X-RAY DIFFRACTION	r_lrange_it	13.402	98.705	42648
X-RAY DIFFRACTION	r_ncsr_local_group_1	0.082	0.05	9828
X-RAY DIFFRACTION	r_ncsr_local_group_2	0.08	0.05	9886
X-RAY DIFFRACTION	r_ncsr_local_group_3	0.081	0.05	9879
X-RAY DIFFRACTION	r_ncsr_local_group_4	0.076	0.05	9950
X-RAY DIFFRACTION	r_ncsr_local_group_5	0.078	0.05	9953
X-RAY DIFFRACTION	r_ncsr_local_group_6	0.071	0.05	10050

Refine LS restraints NCS

Ens-ID	Dom-ID	Auth asym-ID	Refine-ID	Type	Rms dev position (Å)	Weight position
1	1	AAA	X-RAY DIFFRACTION	Local ncs	0.08244	0.05009
1	2	BBB	X-RAY DIFFRACTION	Local ncs	0.08244	0.05009
2	3	AAA	X-RAY DIFFRACTION	Local ncs	0.08009	0.05009
2	4	CCC	X-RAY DIFFRACTION	Local ncs	0.08009	0.05009
3	5	AAA	X-RAY DIFFRACTION	Local ncs	0.08081	0.05009
3	6	DDD	X-RAY DIFFRACTION	Local ncs	0.08081	0.05009
4	7	BBB	X-RAY DIFFRACTION	Local ncs	0.07566	0.05009
4	8	CCC	X-RAY DIFFRACTION	Local ncs	0.07566	0.05009
5	9	BBB	X-RAY DIFFRACTION	Local ncs	0.07788	0.05009
5	10	DDD	X-RAY DIFFRACTION	Local ncs	0.07788	0.05009
6	11	CCC	X-RAY DIFFRACTION	Local ncs	0.07088	0.05009
6	12	DDD	X-RAY DIFFRACTION	Local ncs	0.07088	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
1.957-2.008	0.425	343	0.412	6431	0.412	6802	0.428	0.424	99.5884	0.421
2.008-2.063	0.414	343	0.392	6334	0.393	6691	0.29	0.294	99.7908	0.395
2.063-2.123	0.411	301	0.368	6123	0.37	6436	0.319	0.327	99.8136	0.365
2.123-2.188	0.391	321	0.35	5960	0.352	6295	0.422	0.436	99.7776	0.339
2.188-2.26	0.375	283	0.319	5842	0.322	6142	0.599	0.626	99.7232	0.3
2.26-2.339	0.328	281	0.294	5599	0.296	5892	0.738	0.753	99.7963	0.269
2.339-2.427	0.293	276	0.264	5405	0.265	5694	0.789	0.812	99.7717	0.234
2.427-2.526	0.313	273	0.251	5218	0.254	5507	0.814	0.848	99.7095	0.219
2.526-2.638	0.323	265	0.242	5004	0.246	5288	0.845	0.867	99.6407	0.212
2.638-2.767	0.312	250	0.231	4767	0.235	5038	0.863	0.888	99.5832	0.201
2.767-2.917	0.299	220	0.225	4585	0.228	4814	0.884	0.9	99.813	0.198
2.917-3.094	0.331	213	0.231	4331	0.236	4558	0.862	0.895	99.6928	0.21
3.094-3.307	0.282	184	0.216	4093	0.219	4288	0.891	0.92	99.7435	0.199
3.307-3.572	0.248	218	0.193	3781	0.196	4016	0.92	0.946	99.5767	0.182
3.572-3.913	0.234	179	0.181	3485	0.184	3678	0.939	0.954	99.6194	0.176
3.913-4.374	0.196	139	0.143	3205	0.145	3359	0.954	0.969	99.5534	0.145
4.374-5.05	0.172	159	0.128	2805	0.13	2977	0.967	0.976	99.5633	0.134
5.05-6.183	0.187	111	0.144	2405	0.146	2522	0.961	0.969	99.7621	0.15
6.183-8.736	0.229	104	0.156	1877	0.16	1993	0.953	0.969	99.3979	0.169
8.736-142.548	0.108	39	0.155	1100	0.153	1159	0.988	0.976	98.2744	0.188

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