PDB-7qvr: Structure of E.coli Class 2 L-asparaginase EcAIII, mutant RDM1-37...

ID or keywords:

Entry

Database: PDB / ID: 7qvr

Title

Structure of E.coli Class 2 L-asparaginase EcAIII, mutant RDM1-37 (G206S, R207T, D210S)

Components

Beta-aspartyl-peptidase
Isoaspartyl peptidase

Keywords

HYDROLASE / L-asparaginase / Ntn-hydrolase / EcAIII

Function / homology

Function and homology information

beta-aspartyl-peptidase / asparaginase activity / beta-aspartyl-peptidase activity / protein autoprocessing / hydrolase activity
Similarity search - Function

Biological species

Escherichia coli str. K-12 substr. MG1655 (bacteria)

Escherichia coli (E. coli)

Method

X-RAY DIFFRACTION /

MOLECULAR REPLACEMENT / Resolution: 1.9 Å

Authors

Loch, J.I. / Kadziolka, K. / Jaskolski, M.

Funding support

Poland, 1items

Citation

Journal: Acta Crystallogr D Struct Biol / Year: 2022
Title: Structural and biophysical studies of new L-asparaginase variants: lessons from random mutagenesis of the prototypic Escherichia coli Ntn-amidohydrolase.
Authors: Loch, J.I. / Klonecka, A. / Kadziolka, K. / Bonarek, P. / Barciszewski, J. / Imiolczyk, B. / Brzezinski, K. / Gilski, M. / Jaskolski, M.

History

Deposition	Jan 23, 2022	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Jul 13, 2022	Provider: repository / Type: Initial release
Revision 1.1	Jan 31, 2024	Group: Data collection / Refinement description Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model / struct_ncs_dom_lim Item: _struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id ..._struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id / _struct_ncs_dom_lim.beg_label_comp_id / _struct_ncs_dom_lim.beg_label_seq_id / _struct_ncs_dom_lim.end_auth_comp_id / _struct_ncs_dom_lim.end_label_asym_id / _struct_ncs_dom_lim.end_label_comp_id / _struct_ncs_dom_lim.end_label_seq_id

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	7qvr.cif.gz	220.8 KB	Display	PDBx/mmCIF format
PDB format	pdb7qvr.ent.gz		Display	PDB format
PDBx/mmJSON format	7qvr.json.gz		Tree view	PDBx/mmJSON format
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Validation report

Summary document	7qvr_validation.pdf.gz	456.6 KB	Display	wwPDB validaton report
Full document	7qvr_full_validation.pdf.gz	460.9 KB	Display
Data in XML	7qvr_validation.xml.gz	28.4 KB	Display
Data in CIF	7qvr_validation.cif.gz	41.9 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/qv/7qvr ftp://data.pdbj.org/pub/pdb/validation_reports/qv/7qvr	HTTPS FTP

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Related structure data

Related structure data	7qq8C 7qsfC 7qtcC 7qy6C 7qymC 7qyxC 7r1gC 7r5cC 2zalS 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
Experimental dataset #1	Data reference: 10.18150/R3BTBM / Data set type: diffraction image data

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Links

PDB pages	PDBj / wwPDB / NCBI
Related items in Molecule of the Month	#166 - Oct 2013 Proteasome similarity (1) #80 - Aug 2006 AAA+ Proteases similarity (1) #176 - Aug 2014 Dynein similarity (1) #60 - Dec 2004 Ubiquitin similarity (1)

Deposited unit

AAA: Isoaspartyl peptidase

BBB: Beta-aspartyl-peptidase

CCC: Isoaspartyl peptidase

DDD: Beta-aspartyl-peptidase

hetero molecules

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

1

Idetical with deposited unit
defined by author&software
Evidence: gel filtration

Unit cell

Length a, b, c (Å)	51.224, 74.112, 147.766
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	AAA
2	1	CCC
3	2	BBB
4	2	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	ALA	ALA	ARG	ARG	AAA	A	4 - 157	4 - 157
2	2	1	ALA	ALA	ARG	ARG	CCC	C	4 - 157	4 - 157
3	3	2	THR	THR	GLU	GLU	BBB	B	179 - 313	1 - 135
4	4	2	THR	THR	GLU	GLU	DDD	D	179 - 313	1 - 135

NCS ensembles :

ID	Details
1	Local NCS retraints between domains: 1 2
2	Local NCS retraints between domains: 3 4

#1: Protein	Isoaspartyl peptidase / Beta-aspartyl-peptidase / EcAIII / Isoaspartyl dipeptidase Mass: 19013.773 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli str. K-12 substr. MG1655 (bacteria) Gene: iaaA, spt, ybiK, b0828, JW0812 / Plasmid: pET11d / Production host: Escherichia coli BL21 (bacteria) / Variant (production host): Gold / References: UniProt: P37595, beta-aspartyl-peptidase
#2: Protein	Beta-aspartyl-peptidase Mass: 14374.072 Da / Num. of mol.: 2 / Mutation: G206S, R207T, D210S Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) Gene: iaaA, BE930_09970, C6B13_14600, CDL57_12485, D2183_02875, D9F92_01450, EIA08_01415, ELV10_03530, G5603_02075, GLW94_04880, HH411_000278, HL425_03125, HL601_10320, HLZ20_00445, HND12_01415, HNV94_01710, HVW04_06855 Plasmid: pET11d / Production host: Escherichia coli (E. coli) / References: UniProt: A0A0A1A394, beta-aspartyl-peptidase
#3: Chemical	ChemComp-NA / SODIUM ION Mass: 22.990 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Na
#4: Chemical	ChemComp-CL / CHLORIDE ION Mass: 35.453 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Cl
#5: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 526 / 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: 2.1 Å³/Da / Density % sol: 41.43 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 8.5 / Details: 30% PEG 6000, 0,2 M MgCl2 in 0.1M Tris-HCl pH 8.5

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SEALED TUBE / Type: Agilent SuperNova / Wavelength: 1.54056 Å
Detector	Type: RIGAKU HyPix-6000HE / Detector: PIXEL / Date: Jun 5, 2020
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 1.54056 Å / Relative weight: 1
Reflection	Resolution: 1.9→20.281 Å / Num. obs: 42970 / % possible obs: 95.6 % / Redundancy: 2.6 % / CC_1/2: 0.992 / R_merge(I) obs: 0.079 / R_rim(I) all: 0.097 / Net I/σ(I): 9
Reflection shell	Resolution: 1.9→1.94 Å / R_merge(I) obs: 0.327 / Mean I/σ(I) obs: 2.1 / Num. unique obs: 2773 / CC_1/2: 0.702 / R_rim(I) all: 0.487 / % possible all: 89.8

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0267	refinement
CrysalisPro		data reduction
Aimless		data scaling
PHASER		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 2ZAL

2zal

Resolution: 1.9→20.281 Å / Cor.coef. F_o:F_c: 0.91 / Cor.coef. F_o:F_c free: 0.855 / SU B: 12.748 / SU ML: 0.159 / Cross valid method: FREE R-VALUE / ESU R: 0.201 / ESU R Free: 0.191

	R_factor	Num. reflection	% reflection
R_free	0.2883	1055	2.458 %
R_work	0.2228	41874	-
all	0.224	-	-
obs	-	42929	94.978 %

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: 14.575 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	0.523 Å²	0 Å²	0 Å²
2-	-	0.376 Å²	-0 Å²
3-	-	-	-0.9 Å²

Refinement step

Cycle: LAST / Resolution: 1.9→20.281 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4252	0	4	526	4782

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.011	0.013	4352
X-RAY DIFFRACTION	r_bond_other_d	0.004	0.015	4176
X-RAY DIFFRACTION	r_angle_refined_deg	1.657	1.631	5909
X-RAY DIFFRACTION	r_angle_other_deg	1.439	1.569	9595
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	7.283	5	598
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	30.093	22.183	197
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	14.621	15	700
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	18.365	15	28
X-RAY DIFFRACTION	r_chiral_restr	0.077	0.2	588
X-RAY DIFFRACTION	r_gen_planes_refined	0.008	0.02	5054
X-RAY DIFFRACTION	r_gen_planes_other	0.002	0.02	942
X-RAY DIFFRACTION	r_nbd_refined	0.208	0.2	1016
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.194	0.2	4375
X-RAY DIFFRACTION	r_nbtor_refined	0.159	0.2	2129
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.083	0.2	2050
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.21	0.2	389
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_other	0.119	0.2	1
X-RAY DIFFRACTION	r_metal_ion_refined	0.152	0.2	8
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.273	0.2	27
X-RAY DIFFRACTION	r_nbd_other	0.281	0.2	82
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.217	0.2	23
X-RAY DIFFRACTION	r_xyhbond_nbd_other	0.037	0.2	1
X-RAY DIFFRACTION	r_mcbond_it	0.732	1.173	2356
X-RAY DIFFRACTION	r_mcbond_other	0.729	1.172	2355
X-RAY DIFFRACTION	r_mcangle_it	1.269	1.753	2942
X-RAY DIFFRACTION	r_mcangle_other	1.269	1.754	2943
X-RAY DIFFRACTION	r_scbond_it	0.813	1.323	1996
X-RAY DIFFRACTION	r_scbond_other	0.813	1.32	1994
X-RAY DIFFRACTION	r_scangle_it	1.327	1.932	2959
X-RAY DIFFRACTION	r_scangle_other	1.327	1.932	2959
X-RAY DIFFRACTION	r_lrange_it	3.729	15.003	5038
X-RAY DIFFRACTION	r_lrange_other	3.701	14.978	5030
X-RAY DIFFRACTION	r_ncsr_local_group_1	0.105	0.05	4466
X-RAY DIFFRACTION	r_ncsr_local_group_2	0.097	0.05	3714

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	0.10501	0.05007
1	2	CCC	X-RAY DIFFRACTION	Local ncs	0.10501	0.05007
2	3	BBB	X-RAY DIFFRACTION	Local ncs	0.09715	0.05008
2	4	DDD	X-RAY DIFFRACTION	Local ncs	0.09715	0.05008

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
1.9-1.949	0.366	90	0.334	2918	0.335	3247	0.681	0.729	92.6394	0.316
1.949-2.002	0.379	76	0.305	2941	0.306	3180	0.742	0.789	94.8742	0.286
2.002-2.059	0.359	73	0.276	2974	0.277	3116	0.793	0.839	97.7856	0.26
2.059-2.122	0.347	74	0.271	2886	0.273	2994	0.803	0.836	98.8644	0.259
2.122-2.191	0.336	71	0.273	2835	0.275	2929	0.813	0.846	99.2148	0.255
2.191-2.267	0.322	74	0.242	2740	0.244	2833	0.844	0.881	99.3293	0.225
2.267-2.351	0.299	74	0.224	2655	0.226	2738	0.862	0.899	99.6713	0.205
2.351-2.446	0.337	58	0.224	2593	0.226	2658	0.849	0.898	99.7366	0.205
2.446-2.553	0.291	54	0.241	2472	0.242	2533	0.864	0.882	99.7236	0.222
2.553-2.675	0.303	59	0.236	2347	0.238	2426	0.867	0.888	99.1756	0.219
2.675-2.817	0.25	65	0.218	2232	0.219	2333	0.898	0.908	98.4569	0.2
2.817-2.985	0.311	48	0.219	2097	0.221	2204	0.892	0.907	97.3231	0.204
2.985-3.186	0.295	36	0.205	1970	0.207	2086	0.874	0.911	96.1649	0.192
3.186-3.434	0.312	49	0.195	1779	0.197	1947	0.897	0.933	93.888	0.186
3.434-3.751	0.202	41	0.176	1564	0.177	1813	0.95	0.95	88.5273	0.172
3.751-4.176	0.236	25	0.167	1359	0.169	1635	0.94	0.956	84.6483	0.166
4.176-4.788	0.15	24	0.155	1185	0.155	1483	0.96	0.967	81.5239	0.154
4.788-5.784	0.195	29	0.177	1042	0.178	1281	0.955	0.962	83.6066	0.176
5.784-7.866	0.254	28	0.185	825	0.187	1025	0.929	0.952	83.2195	0.178
7.866-20.281	0.21	7	0.182	460	0.182	684	0.974	0.959	68.2749	0.181

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	0.032	0.0316	-0.0013	0.2276	-0.1425	0.1545	-0.004	0.0023	-0.0271	0.0362	0.0034	0.0141	-0.0133	-0.0034	0.0006	0.0149	-0.0006	0.0145	0.0003	-0.0019	0.0549	2.8029	-4.6749	46.6506
2	0.2027	-0.1268	-0.1026	0.2827	0.134	0.1376	-0.0018	0.0161	-0.0347	0.0252	-0.0128	-0.0209	-0.0383	0.0287	0.0145	0.0392	-0.029	-0.0166	0.0244	0.008	0.0305	17.3248	1.091	43.3456
3	0.2511	-0.0575	-0.185	0.1161	0.0865	0.3393	-0.0347	0.0363	0.0244	-0.0219	0.0059	0.0459	-0.0044	-0.0106	0.0289	0.0173	-0.0131	-0.0162	0.0125	0.0052	0.0409	2.6628	4.1605	13.0748
4	0.0788	-0.0064	0.0641	0.1903	-0.0169	0.2218	-0.003	-0.0131	-0.0151	-0.0349	-0.0173	-0.0197	-0.007	0.0327	0.0204	0.0112	-0.0027	0.0157	0.0198	-0.0016	0.0479	17.6507	0.3874	17.1314

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	ALL	AAA	2 - 162
2	X-RAY DIFFRACTION	1	ALL	AaA	163
3	X-RAY DIFFRACTION	1	ALL	AbA	164 - 350
4	X-RAY DIFFRACTION	2	ALL	BBB	179 - 313
5	X-RAY DIFFRACTION	2	ALL	BaB	314
6	X-RAY DIFFRACTION	2	ALL	BbB	315 - 436
7	X-RAY DIFFRACTION	3	ALL	CCC	4 - 158
8	X-RAY DIFFRACTION	3	ALL	CaC	159
9	X-RAY DIFFRACTION	3	ALL	CbC	160 - 334
10	X-RAY DIFFRACTION	4	ALL	DDD	179 - 313
11	X-RAY DIFFRACTION	4	ALL	DaD	314
12	X-RAY DIFFRACTION	4	ALL	DbD	315 - 410

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