PDB-9ble: Crystal structure of thermostable dienelactone hydrolase. Monocli...

ID or keywords:

Entry

Database: PDB / ID: 9ble

Title

Crystal structure of thermostable dienelactone hydrolase. Monoclinic space group.

Components

Carboxymethylenebutenolidase

Keywords

HYDROLASE / Dienelactone hydrolase / esterase activity / PETase / BHETase / thermostable enzyme / PET depolymerization

Function / homology

: / Dienelactone hydrolase / Dienelactone hydrolase family / Alpha/Beta hydrolase fold / hydrolase activity / Carboxymethylenebutenolidase

Function and homology information

Biological species

Hydrogenobacter thermophilus (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.79 Å

Authors

Muniz, J.R.C. / Almeida, D. / Brito, V. / Ciancaglini, I. / Sandano, A.L.H. / Squina, F. / Garcia, W.

Funding support

Brazil, 25items

Organization	Grant number	Country
Sao Paulo Research Foundation (FAPESP)	2020/15595-3	Brazil
Sao Paulo Research Foundation (FAPESP)	2020/05784-3	Brazil
Sao Paulo Research Foundation (FAPESP)	2022/05731-2	Brazil
Sao Paulo Research Foundation (FAPESP)	2022/08958-8	Brazil
Sao Paulo Research Foundation (FAPESP)	2015/23279-6	Brazil
Sao Paulo Research Foundation (FAPESP)	2019/19360-3	Brazil
Sao Paulo Research Foundation (FAPESP)	2020/11019-8	Brazil
Sao Paulo Research Foundation (FAPESP)	2021/04254-3	Brazil
Sao Paulo Research Foundation (FAPESP)	2022/14112-4	Brazil
Sao Paulo Research Foundation (FAPESP)	2023/04782-5	Brazil
Sao Paulo Research Foundation (FAPESP)	2023/04828-5	Brazil
Sao Paulo Research Foundation (FAPESP)	2023/12398-0	Brazil
Sao Paulo Research Foundation (FAPESP)	2023/04941-6	Brazil
Sao Paulo Research Foundation (FAPESP)	2023/08832-7	Brazil
Brazilian National Council for Scientific and Technological Development (CNPq)	305816/2020-9	Brazil
Brazilian National Council for Scientific and Technological Development (CNPq)	422132/2018-7	Brazil
Brazilian National Council for Scientific and Technological Development (CNPq)	306279/2020-7	Brazil
Brazilian National Council for Scientific and Technological Development (CNPq)	315847/2021-2	Brazil
Brazilian National Council for Scientific and Technological Development (CNPq)	403844/2023-1	Brazil
Brazilian National Council for Scientific and Technological Development (CNPq)	176662/2023-5	Brazil
Brazilian National Council for Scientific and Technological Development (CNPq)	304998/2023-0	Brazil
Brazilian National Council for Scientific and Technological Development (CNPq)	307199/2023-1	Brazil
Other government	88887.625460/2021-00
Other government	88887.600186/2021-00
Other government	88887.518696/2020-00

Citation

Journal: To Be Published
Title: Crystal structure of thermostable dienelactone hydrolase
Authors: Muniz, J.R.C. / Almeida, D. / Brito, V. / Ciancaglini, I. / Sandano, A.L.H. / Squina, F. / Garcia, W.

History

Deposition	Apr 30, 2024	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	May 14, 2025	Provider: repository / Type: Initial release

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	9ble.cif.gz	505.1 KB	Display	PDBx/mmCIF format
PDB format	pdb9ble.ent.gz	318.3 KB	Display	PDB format
PDBx/mmJSON format	9ble.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/bl/9ble ftp://data.pdbj.org/pub/pdb/validation_reports/bl/9ble	HTTPS FTP

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Related structure data

Related structure data	9bld
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: Carboxymethylenebutenolidase

B: Carboxymethylenebutenolidase

C: Carboxymethylenebutenolidase

D: Carboxymethylenebutenolidase

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

1

A: Carboxymethylenebutenolidase

C: Carboxymethylenebutenolidase

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

2

B: Carboxymethylenebutenolidase

D: Carboxymethylenebutenolidase

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

Unit cell

Length a, b, c (Å)	47.474, 142.669, 74.887
Angle α, β, γ (deg.)	90.000, 96.524, 90.000
Int Tables number	4
Space group name H-M	P12₁1

Noncrystallographic symmetry (NCS)

NCS domain:

ID	Ens-ID	Details
1	1	A
2	1	A
3	2	A
4	2	A
5	3	A
6	3	A
7	4	A
8	4	A
9	5	A
10	5	A
11	6	A
12	6	A

NCS domain segments:

Beg auth comp-ID: GLY / Beg label comp-ID: GLY / Auth asym-ID: A / Label asym-ID: A

Dom-ID	Component-ID	Ens-ID	End auth comp-ID	End label comp-ID	Auth seq-ID	Label seq-ID
1	1	1	LEU	LEU	2 - 233	8 - 239
2	1	1	LEU	LEU	2 - 233	8 - 239
3	2	2	LEU	LEU	2 - 233	8 - 239
4	2	2	LEU	LEU	2 - 233	8 - 239
5	3	3	LEU	LEU	2 - 233	8 - 239
6	3	3	LEU	LEU	2 - 233	8 - 239
7	4	4	LEU	LEU	2 - 233	8 - 239
8	4	4	LEU	LEU	2 - 233	8 - 239
9	5	5	THR	THR	2 - 234	8 - 240
10	5	5	THR	THR	2 - 234	8 - 240
11	6	6	LEU	LEU	2 - 233	8 - 239
12	6	6	LEU	LEU	2 - 233	8 - 239

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	Carboxymethylenebutenolidase Mass: 27259.045 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Hydrogenobacter thermophilus (bacteria) Gene: HTH_1817 / Production host: Escherichia coli (E. coli) / References: UniProt: D3DKA9
#2: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 642 / Source method: isolated from a natural source / Formula: H₂O
Has protein modification	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: 2.4 Å³/Da / Density % sol: 48.67 % / Description: long rods
Crystal grow	Temperature: 293 K / Method: vapor diffusion, sitting drop Details: 0.2 M Sodium fluoride, 0.1 M Bis-Tris propane 8.5, 20% PEG 3350

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: LNLS SIRIUS / Beamline: MANACA / Wavelength: 0.97718 Å
Detector	Type: DECTRIS PILATUS 2M / Detector: PIXEL / Date: May 5, 2022
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 0.97718 Å / Relative weight: 1
Reflection	Resolution: 1.53→74.402 Å / Num. obs: 137099 / % possible obs: 92.3 % / Redundancy: 4.7 % / CC_1/2: 0.997 / R_merge(I) obs: 0.121 / R_pim(I) all: 0.062 / R_rim(I) all: 0.137 / Net I/σ(I): 6.6
Reflection shell	Resolution: 1.53→1.556 Å / Redundancy: 5 % / R_merge(I) obs: 4.082 / Mean I/σ(I) obs: 0.5 / Num. unique obs: 7226 / CC_1/2: 0.363 / R_pim(I) all: 1.995 / R_rim(I) all: 4.556 / % possible all: 98.3

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0425	refinement
XDS		data reduction
XDS		data scaling
PHASER		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 1.79→74.402 Å / Cor.coef. F_o:F_c: 0.968 / Cor.coef. F_o:F_c free: 0.953 / SU B: 5.49 / SU ML: 0.084 / Cross valid method: FREE R-VALUE / ESU R: 0.125 / ESU R Free: 0.114
Details: Hydrogens have been added in their riding positions

	R_factor	Num. reflection	% reflection
R_free	0.1903	4200	4.964 %
R_work	0.161	80416	-
all	0.162	-	-
obs	-	84616	91.084 %

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: 26.075 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	2.268 Å²	-0 Å²	0.037 Å²
2-	-	-1.918 Å²	-0 Å²
3-	-	-	-0.35 Å²

Refinement step

Cycle: LAST / Resolution: 1.79→74.402 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	7435	0	0	642	8077

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.008	0.012	7729
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.016	7083
X-RAY DIFFRACTION	r_angle_refined_deg	1.556	1.826	10483
X-RAY DIFFRACTION	r_angle_other_deg	0.547	1.753	16351
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	5.782	5	966
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	6.122	5	36
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	12.126	10	1235
X-RAY DIFFRACTION	r_dihedral_angle_6_deg	17.05	10	368
X-RAY DIFFRACTION	r_chiral_restr	0.079	0.2	1092
X-RAY DIFFRACTION	r_gen_planes_refined	0.007	0.02	9149
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	1845
X-RAY DIFFRACTION	r_nbd_refined	0.212	0.2	1313
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.192	0.2	6042
X-RAY DIFFRACTION	r_nbtor_refined	0.184	0.2	3874
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.08	0.2	3678
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.162	0.2	520
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_other	0.058	0.2	1
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.228	0.2	38
X-RAY DIFFRACTION	r_nbd_other	0.244	0.2	122
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.173	0.2	30
X-RAY DIFFRACTION	r_mcbond_it	1.57	1.586	3786
X-RAY DIFFRACTION	r_mcbond_other	1.57	1.585	3786
X-RAY DIFFRACTION	r_mcangle_it	2.381	2.842	4733
X-RAY DIFFRACTION	r_mcangle_other	2.381	2.842	4734
X-RAY DIFFRACTION	r_scbond_it	2.799	1.945	3943
X-RAY DIFFRACTION	r_scbond_other	2.799	1.945	3944
X-RAY DIFFRACTION	r_scangle_it	4.348	3.407	5735
X-RAY DIFFRACTION	r_scangle_other	4.348	3.407	5736
X-RAY DIFFRACTION	r_lrange_it	5.992	20.528	8807
X-RAY DIFFRACTION	r_lrange_other	5.89	19.594	8658
X-RAY DIFFRACTION	r_ncsr_local_group_1	0.083	0.05	7495
X-RAY DIFFRACTION	r_ncsr_local_group_2	0.08	0.05	7457
X-RAY DIFFRACTION	r_ncsr_local_group_3	0.06	0.05	7565
X-RAY DIFFRACTION	r_ncsr_local_group_4	0.074	0.05	7562
X-RAY DIFFRACTION	r_ncsr_local_group_5	0.083	0.05	7572
X-RAY DIFFRACTION	r_ncsr_local_group_6	0.083	0.05	7448

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	0.08266	0.05009
1	2	A	X-RAY DIFFRACTION	Local ncs	0.08266	0.05009
2	3	A	X-RAY DIFFRACTION	Local ncs	0.07978	0.05009
2	4	A	X-RAY DIFFRACTION	Local ncs	0.07978	0.05009
3	5	A	X-RAY DIFFRACTION	Local ncs	0.05999	0.05009
3	6	A	X-RAY DIFFRACTION	Local ncs	0.05999	0.05009
4	7	A	X-RAY DIFFRACTION	Local ncs	0.07406	0.05009
4	8	A	X-RAY DIFFRACTION	Local ncs	0.07406	0.05009
5	9	A	X-RAY DIFFRACTION	Local ncs	0.08258	0.05009
5	10	A	X-RAY DIFFRACTION	Local ncs	0.08258	0.05009
6	11	A	X-RAY DIFFRACTION	Local ncs	0.08293	0.05009
6	12	A	X-RAY DIFFRACTION	Local ncs	0.08293	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
1.79-1.836	0.288	317	0.277	6476	0.278	6864	0.933	0.938	98.9656	0.271
1.836-1.887	0.263	278	0.249	5203	0.25	6639	0.949	0.953	82.5576	0.238
1.887-1.941	0.238	231	0.234	4927	0.234	6501	0.956	0.959	79.3416	0.221
1.941-2.001	0.2	283	0.189	4944	0.19	6332	0.971	0.974	82.549	0.171
2.001-2.067	0.199	252	0.172	5110	0.173	6139	0.971	0.98	87.3432	0.159
2.067-2.139	0.199	273	0.161	4950	0.163	5904	0.973	0.983	88.4654	0.149
2.139-2.22	0.197	252	0.154	5420	0.156	5695	0.976	0.986	99.5961	0.144
2.22-2.31	0.163	214	0.146	4103	0.147	5478	0.982	0.986	78.8061	0.136
2.31-2.413	0.189	235	0.137	5040	0.139	5292	0.981	0.988	99.6788	0.131
2.413-2.531	0.177	260	0.143	4766	0.145	5035	0.98	0.987	99.8213	0.138
2.531-2.667	0.192	239	0.152	4081	0.155	4818	0.978	0.986	89.6638	0.15
2.667-2.829	0.208	224	0.164	4039	0.166	4534	0.971	0.983	94.0229	0.165
2.829-3.024	0.197	213	0.159	4038	0.161	4257	0.976	0.985	99.8591	0.166
3.024-3.266	0.193	189	0.154	3644	0.156	3984	0.977	0.986	96.2098	0.168
3.266-3.577	0.181	169	0.16	3106	0.161	3660	0.979	0.985	89.4809	0.179
3.577-3.998	0.174	139	0.148	2562	0.149	3328	0.983	0.988	81.1599	0.17
3.998-4.614	0.147	144	0.127	2778	0.128	2922	0.988	0.99	100	0.156
4.614-5.646	0.174	132	0.139	2334	0.141	2466	0.982	0.99	100	0.172
5.646-7.961	0.19	107	0.168	1843	0.17	1950	0.979	0.984	100	0.202
7.961-74.402	0.217	48	0.199	1037	0.2	1085	0.965	0.967	100	0.238

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	0.6291	0.4074	0.4099	1.8737	0.0391	1.2364	-0.0076	-0.0791	-0.0428	0.0921	-0.0156	-0.1912	0.0905	-0.0124	0.0232	0.1262	0.009	-0.0095	0.0739	0.0184	0.0893	-16.7901	32.1169	-41.4019
2	0.8037	1.0382	-0.9242	2.1696	-1.122	1.4462	-0.0645	0.1028	-0.13	-0.1078	0.1208	-0.0345	0.1172	-0.1318	-0.0563	0.1138	0.0008	0.0035	0.0146	-0.0079	0.1701	-22.7457	27.8313	-50.2867
3	0.0528	0.2478	0.0694	1.2941	0.1866	0.8068	-0.0428	-0.0263	0.0093	-0.1203	-0.0508	0.064	-0.0365	-0.0614	0.0936	0.1128	0.0342	0.0148	0.0995	-0.0165	0.0826	-27.9136	39.566	-47.6044
4	0.3552	0.4242	0.0898	1.2363	0.1641	0.2363	-0.0586	-0.0942	0.0355	-0.1639	0.0354	0.0031	-0.1916	0.0172	0.0232	0.1626	-0.0227	-0.0091	0.0606	-0.0227	0.0594	-19.1356	49.4313	-50.8644
5	2.6318	-2.5157	-2.7024	7.0392	4.1643	4.2725	0.0483	0.1237	0.0437	-0.1909	-0.076	-0.0129	-0.1447	-0.2157	0.0277	0.0875	-0.0051	-0.0085	0.0822	0.0336	0.1046	-0.5654	16.7251	3.7276
6	9.0639	1.1276	-10.9017	0.8785	0.6992	18.8369	-0.5033	1.1005	-0.1586	-0.2994	-0.0544	0.1008	-0.0473	-1.8755	0.5577	0.1389	-0.03	-0.1531	0.2259	0.0672	0.4668	-7.1174	4.2033	-7.3264
7	0.1983	0.0058	0.1802	1.4413	-0.0614	0.4696	-0.0346	0.0511	0.0407	0.0266	0.0573	-0.0773	-0.0706	0.0305	-0.0227	0.0685	-0.0416	0.0204	0.0649	0.0023	0.1254	5.3087	9.441	8.5626
8	0.329	0.0903	0.0549	1.0042	-0.1642	0.0656	-0.0087	0.031	0.0159	0.0762	0.0351	-0.0401	0.002	-0.0293	-0.0263	0.0522	-0.0347	0.0196	0.0877	-0.0114	0.1282	3.4988	-4.5463	12.6166
9	0.074	0.2678	0.0716	1.265	-0.0983	0.9419	-0.0373	0.0239	-0.044	0.0788	0.0144	-0.2106	-0.1618	-0.0062	0.0229	0.203	-0.0595	-0.0307	0.0512	-0.0024	0.059	-14.6492	8.9492	-32.8016
10	0.5123	-0.1899	0.7173	1.2478	0.8885	2.279	-0.1079	0.1005	-0.0598	0.1934	-0.0122	0.2634	-0.1437	0.1156	0.1201	0.2508	0.0023	0.0442	0.0374	0.0119	0.0825	-27.1513	9.789	-30.0303
11	0.767	-0.082	-0.063	1.1475	-0.3688	0.1293	-0.0499	0.1366	-0.0161	0.4166	0.0149	-0.0952	-0.1275	-0.0214	0.035	0.2472	-0.0138	-0.0362	0.0278	-0.0029	0.0361	-17.7015	-4.0057	-20.4248
12	7.5575	-1.7323	1.7409	1.9105	-0.5817	1.8621	0.105	0.1854	-0.175	-0.0665	-0.0472	-0.0762	0.1443	0.0482	-0.0578	0.1723	-0.0084	-0.0072	0.0668	-0.0194	0.0861	-17.4741	-9.4607	-33.8121
13	9.3992	3.0106	1.5538	4.6257	-0.6015	0.5942	-0.0362	-0.2737	0.0557	-0.0671	-0.0137	0.0152	0.0326	-0.0633	0.05	0.1515	-0.0089	0.0399	0.0817	0.0055	0.1269	-3.5224	22.5613	-0.6588
14	0.4089	0.05	0.2547	1.8862	0.1509	2.1663	0.0517	-0.0436	-0.0434	-0.1143	-0.0376	-0.0764	0.1262	0.0516	-0.0141	0.0828	0.0228	0.0067	0.0767	0.0036	0.1099	1.0709	33.2602	-6.5791
15	1.1293	0.2756	0.8223	1.2573	-0.6024	9.2656	0.016	0.0305	-0.1544	-0.0892	0.1334	0.3198	0.1705	-0.218	-0.1495	0.0917	-0.0196	-0.0322	0.0561	0.007	0.1682	-10.9519	28.0578	-6.3132
16	0.0255	-0.1866	0.0695	1.5664	-0.6067	0.4879	0.0232	-0.0138	-0.0073	-0.1016	0.0767	0.1926	0.0455	0.0196	-0.0999	0.0581	0.0034	0.0109	0.0844	0.0034	0.1196	-7.1905	45.5426	-10.3208

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	ALL	A	2 - 69
2	X-RAY DIFFRACTION	2	ALL	A	70 - 103
3	X-RAY DIFFRACTION	3	ALL	A	104 - 158
4	X-RAY DIFFRACTION	4	ALL	A	159 - 234

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