PDB-5h8l: Crystal structure of Medicago truncatula N-carbamoylputrescine am...

ID or keywords:

Entry

Database: PDB / ID: 5h8l

Title

Crystal structure of Medicago truncatula N-carbamoylputrescine amidohydrolase (MtCPA) C158S mutant in complex with putrescine

Components

N-carbamoylputrescine amidohydrolase

Keywords

HYDROLASE / amidase / helical octamer / alpha-beta-beta-alpha sandwich fold

Function / homology

Function and homology information

N-carbamoylputrescine amidase / N-carbamoylputrescine amidase activity / putrescine biosynthetic process from arginine
Similarity search - Function

Biological species

Medicago truncatula (barrel medic)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.29 Å

Authors

Sekula, B. / Ruszkowski, M. / Malinska, M. / Dauter, Z.

Funding support

Poland, 1items

Citation

Journal: Front Plant Sci / Year: 2016
Title: Structural Investigations of N-carbamoylputrescine Amidohydrolase from Medicago truncatula: Insights into the Ultimate Step of Putrescine Biosynthesis in Plants.
Authors: Sekula, B. / Ruszkowski, M. / Malinska, M. / Dauter, Z.

History

Deposition	Dec 23, 2015	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Apr 20, 2016	Provider: repository / Type: Initial release
Revision 1.1	Sep 27, 2023	Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / pdbx_prerelease_seq / pdbx_struct_oper_list Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_oper_list.symmetry_operation

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	5h8l.cif.gz	1.9 MB	Display	PDBx/mmCIF format
PDB format	pdb5h8l.ent.gz	1.6 MB	Display	PDB format
PDBx/mmJSON format	5h8l.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/h8/5h8l ftp://data.pdbj.org/pub/pdb/validation_reports/h8/5h8l	HTTPS FTP

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Related structure data

Related structure data	5h8iC 5h8jC 5h8kC 1erzS C: citing same article (ref.) S: Starting model for refinement
Similar structure data	5h8i-1 5h8j-1 5h8k-1 5h8j-2 5h8k-2 5h8i-2 5c16-d 6ybb-d Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) Search similar-shape structures by Omokage search (details) 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: N-carbamoylputrescine amidohydrolase

B: N-carbamoylputrescine amidohydrolase

C: N-carbamoylputrescine amidohydrolase

D: N-carbamoylputrescine amidohydrolase

E: N-carbamoylputrescine amidohydrolase

F: N-carbamoylputrescine amidohydrolase

G: N-carbamoylputrescine amidohydrolase

H: N-carbamoylputrescine amidohydrolase

I: N-carbamoylputrescine amidohydrolase

J: N-carbamoylputrescine amidohydrolase

K: N-carbamoylputrescine amidohydrolase

L: N-carbamoylputrescine amidohydrolase

M: N-carbamoylputrescine amidohydrolase

N: N-carbamoylputrescine amidohydrolase

O: N-carbamoylputrescine amidohydrolase

P: N-carbamoylputrescine amidohydrolase

hetero molecules

551 kDa, 16 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: N-carbamoylputrescine amidohydrolase

B: N-carbamoylputrescine amidohydrolase

C: N-carbamoylputrescine amidohydrolase

D: N-carbamoylputrescine amidohydrolase

E: N-carbamoylputrescine amidohydrolase

F: N-carbamoylputrescine amidohydrolase

G: N-carbamoylputrescine amidohydrolase

H: N-carbamoylputrescine amidohydrolase

hetero molecules

defined by author&software
275 kDa, 8 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

I: N-carbamoylputrescine amidohydrolase

J: N-carbamoylputrescine amidohydrolase

K: N-carbamoylputrescine amidohydrolase

L: N-carbamoylputrescine amidohydrolase

M: N-carbamoylputrescine amidohydrolase

N: N-carbamoylputrescine amidohydrolase

O: N-carbamoylputrescine amidohydrolase

P: N-carbamoylputrescine amidohydrolase

hetero molecules

defined by author&software
276 kDa, 8 polymers
Search similar-shape structures of this assembly by Omokage search (details)

Unit cell

Length a, b, c (Å)	152.210, 211.082, 208.613
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	18
Space group name H-M	P2₁2₁2

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Protein , 1 types, 16 molecules ABCDEFGHIJKLMNOP

#1: Protein	N-carbamoylputrescine amidohydrolase Mass: 34114.594 Da / Num. of mol.: 16 / Mutation: C158S Source method: isolated from a genetically manipulated source Source: (gene. exp.) Medicago truncatula (barrel medic) / Gene: MTR_2g086600 / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21 GOLD (DE3) / References: UniProt: G7ITU5

#1: Protein

N-carbamoylputrescine amidohydrolase

Mass: 34114.594 Da / Num. of mol.: 16 / Mutation: C158S
Source method: isolated from a genetically manipulated source
Source: (gene. exp.)

Medicago truncatula (barrel medic) / Gene: MTR_2g086600 / Production host:

Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21 GOLD (DE3) / References: UniProt: G7ITU5

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Non-polymers , 5 types, 3767 molecules

#2: Chemical	... ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL Mass: 92.094 Da / Num. of mol.: 30 / Source method: obtained synthetically / Formula: C₃H₈O₃
#3: Chemical	ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL Mass: 62.068 Da / Num. of mol.: 17 / Source method: obtained synthetically / Formula: C₂H₆O₂
#4: Chemical	ChemComp-PUT / 1,4-DIAMINOBUTANE / PUTRESCINE Mass: 88.151 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C₄H₁₂N₂
#5: Chemical	ChemComp-PEG / DI(HYDROXYETHYL)ETHER Mass: 106.120 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C₄H₁₀O₃
#6: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 3705 / Source method: isolated from a natural source / Formula: H₂O

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Experiment

Experiment	Method: X-RAY DIFFRACTION

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Sample preparation

Crystal	Density Matthews: 3.07 Å³/Da / Density % sol: 59.93 %
Crystal grow	Temperature: 292 K / Method: vapor diffusion, hanging drop / pH: 7 / Details: 20% PEG3350, 8% Tacsimate at pH 7.0, 10% glycerol

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Data collection

Diffraction	Mean temperature: 100 K
Diffraction source	Source: SYNCHROTRON / Site: APS / Beamline: 22-ID / Wavelength: 1 Å
Detector	Type: RAYONIX MX300-HS / Detector: CCD / Date: Aug 23, 2015
Radiation	Monochromator: Si (111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 1 Å / Relative weight: 1
Reflection	Resolution: 2.29→40 Å / Num. obs: 294391 / % possible obs: 97.5 % / Observed criterion σ(I): -3 / Redundancy: 3.79 % / R_merge(I) obs: 0.118 / Net I/σ(I): 8.68
Reflection shell	Resolution: 2.29→2.42 Å / Redundancy: 3.78 % / R_merge(I) obs: 0.615 / Mean I/σ(I) obs: 1.97 / % possible all: 88

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0135	refinement
XSCALE		data scaling
XDS		data reduction
BALBES		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 1ERZ

1erz

Resolution: 2.29→39.53 Å / Cor.coef. F_o:F_c: 0.974 / Cor.coef. F_o:F_c free: 0.952 / SU B: 12.384 / SU ML: 0.154 / Cross valid method: FREE R-VALUE / ESU R: 0.206 / ESU R Free: 0.18 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS

	R_factor	Num. reflection	% reflection	Selection details
R_free	0.206	2944	1 %	RANDOM
R_work	0.154	-	-	-
obs	0.155	291447	97.5 %	-

Solvent computation

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

Displacement parameters

B_iso mean: 46.7 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	-1.42 Å²	0 Å²	0 Å²
2-	-	1.64 Å²	0 Å²
3-	-	-	-0.22 Å²

Refinement step

Cycle: LAST / Resolution: 2.29→39.53 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	37559	0	341	3705	41605

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.017	0.019	38795
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.02	37007
X-RAY DIFFRACTION	r_angle_refined_deg	1.6	1.9	52392
X-RAY DIFFRACTION	r_angle_other_deg	0.796	3	85077
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	5.973	5	4742
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	38.87	24.082	1884
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	14.015	15	6508
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	17.926	15	240
X-RAY DIFFRACTION	r_chiral_restr	0.118	0.2	5631
X-RAY DIFFRACTION	r_gen_planes_refined	0.02	0.021	44052
X-RAY DIFFRACTION	r_gen_planes_other	0.015	0.02	9214
X-RAY DIFFRACTION	r_nbd_refined
X-RAY DIFFRACTION	r_nbd_other
X-RAY DIFFRACTION	r_nbtor_refined
X-RAY DIFFRACTION	r_nbtor_other
X-RAY DIFFRACTION	r_xyhbond_nbd_refined
X-RAY DIFFRACTION	r_xyhbond_nbd_other
X-RAY DIFFRACTION	r_metal_ion_refined
X-RAY DIFFRACTION	r_metal_ion_other
X-RAY DIFFRACTION	r_symmetry_vdw_refined
X-RAY DIFFRACTION	r_symmetry_vdw_other
X-RAY DIFFRACTION	r_symmetry_hbond_refined
X-RAY DIFFRACTION	r_symmetry_hbond_other
X-RAY DIFFRACTION	r_symmetry_metal_ion_refined
X-RAY DIFFRACTION	r_symmetry_metal_ion_other
X-RAY DIFFRACTION	r_mcbond_it	1.667	1.596	18938
X-RAY DIFFRACTION	r_mcbond_other	1.667	1.596	18937
X-RAY DIFFRACTION	r_mcangle_it	2.568	2.381	23639
X-RAY DIFFRACTION	r_mcangle_other	2.568	2.381	23640
X-RAY DIFFRACTION	r_scbond_it	2.702	1.89	19857
X-RAY DIFFRACTION	r_scbond_other	2.697	1.89	19857
X-RAY DIFFRACTION	r_scangle_it
X-RAY DIFFRACTION	r_scangle_other	3.973	2.697	28737
X-RAY DIFFRACTION	r_long_range_B_refined	8.404	14.919	46601
X-RAY DIFFRACTION	r_long_range_B_other	8.404	14.921	46602
X-RAY DIFFRACTION	r_rigid_bond_restr
X-RAY DIFFRACTION	r_sphericity_free
X-RAY DIFFRACTION	r_sphericity_bonded

LS refinement shell

Resolution: 2.29→2.35 Å

	R_factor	Num. reflection	% reflection
R_free	0.291	163	-
R_work	0.259	16212	-
obs	-	-	74.05 %

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	1.641	-0.3232	-0.6191	2.8097	1.782	2.6896	-0.0862	-0.433	0.0891	0.3296	0.0398	-0.0382	-0.0509	0.0151	0.0464	0.3202	0.02	-0.005	0.207	-0.1518	0.1897	38.526	92.466	-13.41
2	2.0526	0.1497	-0.0207	1.4715	0.3129	1.0149	-0.022	-0.2296	0.1282	0.016	0.0502	-0.2591	-0.008	0.1737	-0.0282	0.2669	-0.019	0.0336	0.0753	-0.06	0.1303	57.849	74.732	-34.189
3	0.7549	0.1586	0.1107	2.3103	0.0406	1.3509	-0.0203	0.0293	0.2094	-0.3109	-0.0377	0.3328	-0.1503	-0.0708	0.0579	0.3383	0.0473	-0.0734	0.0109	-0.0045	0.1069	23.871	74.163	-53.982
4	0.7	0.2522	0.1618	1.8567	0.2883	1.2899	0.016	0.1065	-0.0959	-0.3231	0.0445	-0.059	0.0458	0.0663	-0.0604	0.3829	0.0386	-0.0068	0.0237	-0.0198	0.0191	35.851	43.87	-61.099
5	1.6338	-0.0619	0.3884	1.3309	-0.1286	1.2139	0.0358	-0.2593	-0.1163	0.0973	-0.0733	0.2965	0.0527	-0.193	0.0375	0.2948	-0.0443	-0.0056	0.0859	0.0152	0.1414	10.836	35.243	-32.467
6	1.4323	-0.0872	0.4113	2.157	-0.9391	1.4477	0.1607	-0.2333	-0.4013	0.0793	-0.136	-0.0756	0.2077	0.0759	-0.0247	0.4062	-0.0321	-0.0675	0.0866	0.1362	0.2332	35.082	13.269	-25.938
7	1.5174	-0.3063	0.0948	1.0591	-0.1254	2.044	0.0186	-0.4239	0.1175	0.2828	-0.0385	-0.1744	-0.3407	0.1796	0.0199	0.7562	-0.175	-0.1051	0.4851	0.1115	0.2733	43.517	40.832	0.041
8	4.1422	-0.2644	0.3821	1.2199	0.6512	1.555	0.1065	-0.155	0.0216	0.1018	0.0615	-0.7473	-0.1331	0.6236	-0.168	0.58	-0.1451	-0.1842	0.8686	0.2246	0.7119	74.228	29.71	-8.406
9	1.4892	-0.2938	-0.6795	2.7795	1.9669	2.837	-0.0174	-0.2881	0.0305	0.2029	0.0758	-0.0007	-0.1019	0.1175	-0.0584	0.3325	-0.0307	0.0317	0.09	-0.0736	0.1594	38.884	88.267	90.558
10	1.8428	0.1814	-0.199	1.3514	0.2517	1.1775	-0.0003	-0.1455	0.0969	0.0072	0.0372	-0.236	-0.0328	0.2132	-0.0369	0.3037	-0.0495	0.0502	0.0687	-0.0572	0.1379	58.115	70.065	69.978
11	0.7443	0.2298	0.0475	2.2084	-0.0404	1.342	-0.0133	0.0789	0.1956	-0.3312	0.0087	0.3188	-0.2256	-0.0757	0.0047	0.4084	0.0511	-0.0595	0.0209	0.0087	0.1039	24.12	69.681	50.195
12	0.631	0.2372	0.076	2.0399	0.2455	1.3532	-0.0404	0.1289	-0.0811	-0.3616	0.0846	-0.0724	-0.0056	0.0605	-0.0442	0.3606	0.0113	0.0002	0.0399	-0.0269	0.0186	35.785	39.212	43.298
13	1.3124	-0.156	0.5519	1.2572	-0.1398	1.2079	-0.0627	-0.1441	-0.0781	0.0874	0.0287	0.3673	0.0525	-0.1847	0.034	0.2642	0.0195	0.0257	0.0494	0.0028	0.1653	10.397	30.931	71.74
14	1.0155	-0.1614	0.4453	1.8579	-0.7375	1.1765	0.0617	-0.0264	-0.2274	0.1102	-0.0267	0.0454	0.1409	0.0375	-0.035	0.342	0.0291	0.0064	0.0063	0.0184	0.1243	34.44	8.865	78.647
15	1.5489	-0.3192	0.0925	1.0889	0.155	1.6659	-0.1381	-0.2764	0.207	0.2992	0.0982	-0.1137	-0.2007	0.0577	0.0399	0.5621	0.043	-0.092	0.1682	-0.0108	0.1531	42.648	36.657	104.664
16	4.4135	-0.6959	0.3101	0.9692	0.6026	1.6726	-0.0163	-0.0784	0.2504	0.1382	0.2022	-0.5178	-0.0906	0.604	-0.186	0.457	0.0367	-0.183	0.4714	-0.0319	0.4357	73.486	25.764	96.671

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	A	1 - 303
2	X-RAY DIFFRACTION	2	B	1 - 303
3	X-RAY DIFFRACTION	3	C	1 - 303
4	X-RAY DIFFRACTION	4	D	1 - 303
5	X-RAY DIFFRACTION	5	E	1 - 303
6	X-RAY DIFFRACTION	6	F	1 - 303
7	X-RAY DIFFRACTION	7	G	1 - 303
8	X-RAY DIFFRACTION	8	H	1 - 303
9	X-RAY DIFFRACTION	9	I	1 - 303
10	X-RAY DIFFRACTION	10	J	1 - 303
11	X-RAY DIFFRACTION	11	K	1 - 303
12	X-RAY DIFFRACTION	12	L	1 - 303
13	X-RAY DIFFRACTION	13	M	1 - 303
14	X-RAY DIFFRACTION	14	N	1 - 303
15	X-RAY DIFFRACTION	15	O	1 - 303
16	X-RAY DIFFRACTION	16	P	1 - 303

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