PDB-8ruf: Crystal structure of Rhizobium etli L-asparaginase ReAV D187A mutant

ID or keywords:

Entry

Database: PDB / ID: 8ruf

Title

Crystal structure of Rhizobium etli L-asparaginase ReAV D187A mutant

Components

L-asparaginase II protein

Keywords

HYDROLASE / Rhizobium etli / amidohydrolases / l-asparaginases / zinc-binding proteins / site-directed mutagenesis

Function / homology

L-asparaginase II / L-asparaginase II / metal ion binding / DI(HYDROXYETHYL)ETHER / L-asparaginase II protein

Function and homology information

Biological species

Rhizobium etli (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.6 Å

Authors

Pokrywka, K. / Grzechowiak, M. / Sliwiak, J. / Worsztynowicz, P. / Loch, J.I. / Ruszkowski, M. / Gilski, M. / Jaskolski, M.

Funding support

Poland, 1items

Citation

Journal: Front Chem / Year: 2024
Title: Probing the active site of Class 3 L-asparaginase by mutagenesis. I. Tinkering with the zinc coordination site of ReAV.
Authors: Pokrywka, K. / Grzechowiak, M. / Sliwiak, J. / Worsztynowicz, P. / Loch, J.I. / Ruszkowski, M. / Gilski, M. / Jaskolski, M.

History

Deposition	Jan 30, 2024	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Apr 17, 2024	Provider: repository / Type: Initial release
Revision 1.1	May 1, 2024	Group: Database references / Category: citation Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	8ruf.cif.gz	577.2 KB	Display	PDBx/mmCIF format
PDB format	pdb8ruf.ent.gz	469.5 KB	Display	PDB format
PDBx/mmJSON format	8ruf.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/ru/8ruf ftp://data.pdbj.org/pub/pdb/validation_reports/ru/8ruf	HTTPS FTP

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Related structure data

Related structure data	8ruaC 8rudC 8rueC 8rugC 7os3 7os5 7os6 7ou1 7oz6 8cly 8clz 8col 8ori 8osw C: citing same article (ref.)
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/BWIJSJ / Data set type: diffraction image data / Metadata reference: 10.18150/BWIJSJ

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Links

PDB pages	PDBj / wwPDB / NCBI

Deposited unit

A: L-asparaginase II protein

B: L-asparaginase II protein

C: L-asparaginase II protein

D: L-asparaginase II protein

hetero molecules

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

1

A: L-asparaginase II protein

B: L-asparaginase II protein

hetero molecules

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

2

C: L-asparaginase II protein

D: L-asparaginase II protein

hetero molecules

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

Unit cell

Length a, b, c (Å)	77.715, 91.260, 114.197
Angle α, β, γ (deg.)	90.000, 96.950, 90.000
Int Tables number	4
Space group name H-M	P12₁1
Space group name Hall	P2yb
Symmetry operation	#1: x,y,z #2: -x,y+1/2,-z

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Protein , 1 types, 4 molecules ABCD

#1: Protein	L-asparaginase II protein Mass: 39510.859 Da / Num. of mol.: 4 / Mutation: D187A Source method: isolated from a genetically manipulated source Details: point mutation D187A / Source: (gene. exp.) Rhizobium etli (bacteria) / Gene: ansA, RHE_PE00350 / Production host: Escherichia coli (E. coli) / References: UniProt: Q2K0Z2

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Non-polymers , 8 types, 1576 molecules

#2: Chemical	ChemComp-SO4 / SULFATE ION Mass: 96.063 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: SO₄
#3: Chemical	... ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL Mass: 62.068 Da / Num. of mol.: 21 / Source method: obtained synthetically / Formula: C₂H₆O₂
#4: Chemical	ChemComp-ZN / ZINC ION Mass: 65.409 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Zn / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical	ChemComp-CL / CHLORIDE ION Mass: 35.453 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Cl
#6: Chemical	ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL Mass: 92.094 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C₃H₈O₃
#7: Chemical	ChemComp-TRS / 2-AMINO-2-HYDROXYMETHYL-PROPANE-1,3-DIOL / TRIS BUFFER Mass: 122.143 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₄H₁₂NO₃ / Comment: pH buffer*YM
#8: Chemical	ChemComp-PEG / DI(HYDROXYETHYL)ETHER Mass: 106.120 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₄H₁₀O₃
#9: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 1534 / 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: 2.54 Å³/Da / Density % sol: 51.64 %
Crystal grow	Temperature: 292 K / Method: vapor diffusion, hanging drop / pH: 9 / Details: 22% PEG 3350, 0.2 M Li2SO4, 0.1 M Bicine pH 9.0

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: PETRA III, EMBL c/o DESY / Beamline: P13 (MX1) / Wavelength: 0.8265 Å
Detector	Type: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Nov 21, 2022
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 0.8265 Å / Relative weight: 1
Reflection	Resolution: 1.6→113.36 Å / Num. obs: 208980 / % possible obs: 99.9 % / Redundancy: 7 % / B_iso Wilson estimate: 18.2 Å² / CC_1/2: 0.999 / R_merge(I) obs: 0.119 / R_rim(I) all: 0.129 / Net I/σ(I): 12.66
Reflection shell	Resolution: 1.6→1.69 Å / R_merge(I) obs: 1.286 / Mean I/σ(I) obs: 1.63 / Num. unique obs: 33605 / CC_1/2: 0.675 / R_rim(I) all: 1.388

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Processing

Software

Name	Version	Classification
PHENIX	1.20.1_4487	refinement
XDS		data reduction
XSCALE		data scaling
PHASER		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 1.6→48.57 Å / SU ML: 0.1642 / Cross valid method: FREE R-VALUE / σ(F): 1.36 / Phase error: 20.0335
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Details: Hydrogen atoms were added at riding position.

	R_factor	Num. reflection	% reflection
R_free	0.1841	2087	1 %
R_work	0.1576	206802	-
obs	0.1578	208889	99.85 %

Solvent computation

Shrinkage radii: 0.9 Å / VDW probe radii: 1.1 Å / Solvent model: FLAT BULK SOLVENT MODEL

Displacement parameters

B_iso mean: 24.26 Å²

Refinement step

Cycle: LAST / Resolution: 1.6→48.57 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	10340	0	172	1534	12046

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.0099	11145
X-RAY DIFFRACTION	f_angle_d	1.0823	15118
X-RAY DIFFRACTION	f_chiral_restr	0.0674	1681
X-RAY DIFFRACTION	f_plane_restr	0.0105	2000
X-RAY DIFFRACTION	f_dihedral_angle_d	13.2792	4236

LS refinement shell

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Refine-ID	% reflection obs (%)
1.6-1.63	0.2526	137	0.264	13585	X-RAY DIFFRACTION	98.73
1.63-1.68	0.28	139	0.2471	13723	X-RAY DIFFRACTION	99.96
1.68-1.72	0.2599	139	0.2326	13799	X-RAY DIFFRACTION	99.98
1.72-1.77	0.3	139	0.2182	13730	X-RAY DIFFRACTION	99.99
1.77-1.83	0.2409	139	0.2011	13776	X-RAY DIFFRACTION	99.99
1.83-1.89	0.2088	139	0.1869	13785	X-RAY DIFFRACTION	99.98
1.89-1.97	0.2122	139	0.1718	13745	X-RAY DIFFRACTION	99.97
1.97-2.06	0.1883	139	0.1533	13771	X-RAY DIFFRACTION	99.96
2.06-2.17	0.2014	138	0.1517	13774	X-RAY DIFFRACTION	99.94
2.17-2.3	0.1871	140	0.148	13779	X-RAY DIFFRACTION	99.91
2.3-2.48	0.1769	139	0.1483	13803	X-RAY DIFFRACTION	99.95
2.48-2.73	0.1854	139	0.1479	13809	X-RAY DIFFRACTION	99.97
2.73-3.13	0.1674	140	0.1504	13858	X-RAY DIFFRACTION	99.94
3.13-3.94	0.1533	141	0.1352	13873	X-RAY DIFFRACTION	99.96
3.94-48.57	0.1511	140	0.1385	13992	X-RAY DIFFRACTION	99.49

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	0.338175643423	-0.711445251689	-0.263933542648	2.77158197336	-0.0830315141003	0.779045549093	-0.0678861306677	-0.0834632466202	0.0112124857303	0.211110692345	0.0545410085533	0.0327035692414	0.0587204668893	0.209625115945	0.022179707437	0.117136579787	0.0188209389692	-0.0178147495149	0.175037628323	0.000871061181562	0.170238730604	22.614	45.937	65.172
2	1.83961175448	-0.326404421493	-0.16672377834	1.59079269346	0.379601161559	1.53144243357	-0.00105973639522	0.0534190900724	0.0171659693039	-0.114745444743	-0.029065582905	0.0735205201554	0.00364698997335	-0.0455967989198	0.018414292409	0.136717352375	0.0339493373257	-0.00135045627769	0.133270931654	0.0028544944686	0.140583507431	28.556	29.948	39.112
3	0.537723768417	-0.0104685062843	-0.302071866439	1.23414206666	0.314785278459	1.40831749927	-0.0645691459949	-0.142782243589	-0.023292715719	0.0280583892177	0.0557255771597	-0.192392932615	0.198691431661	0.38330518962	0.0275801049851	0.134751858502	0.0912818071577	-0.0138068111746	0.236814934655	0.00293957758409	0.186790488597	34.831	33.315	53.496
4	1.45237524242	-0.671847862611	-0.0414574080776	1.21504865458	-0.150335119867	0.968529327468	-0.0512681178647	-0.0465308485007	0.129402508166	0.0542960139844	0.0987969861603	-0.150340925044	-0.0778113895515	0.264186859917	-0.00042587915909	0.104442425754	-0.0147133451751	-0.0147537925269	0.196949206239	-0.0233632283726	0.189145846705	29.086	51.8	57.821
5	5.48621853109	-6.41604621259	-2.32339277681	7.60459673781	2.37621927223	1.39434506026	-0.244624364592	-0.247882540887	-0.13662840096	0.289673022095	0.221109793045	-0.0715145371681	0.152241008424	0.125079739357	-0.0322893684219	0.123767291275	0.000103212679312	-0.00505764593978	0.133243997352	0.00962698605423	0.169239651693	6.576	41.101	62.753
6	0.723821596917	-1.05191551463	1.06211864386	3.94736162605	-0.174076236108	2.49129930633	-0.0614833638733	-0.148227695832	-0.10151693026	0.226118418344	0.0581325976209	0.22088941173	-0.100602715637	-0.255530532331	-0.00193569693362	0.132837198021	0.0220670935343	0.00244238360459	0.134152188489	-0.0149532232468	0.153639444869	-3.717	54.24	67.073
7	0.671853866093	-0.240137394898	-0.22003747194	0.748414474212	0.0745606887035	1.54820043836	0.0459016136849	0.0650085171325	0.0404838679614	-0.0927924924786	-0.0688742168168	-0.017869967164	-0.0437653127479	-0.0207168152378	0.0219686775409	0.130587527301	0.0127820454067	-0.0109678770532	0.0991959267782	0.00419503028018	0.161685077409	-5.208	64.641	40.22
8	3.66814388969	-2.34627908015	-1.68379629585	4.61052380349	2.04363057493	3.42154096718	0.18834519281	0.181409486754	-0.134766677113	-0.293842849926	-0.165139191752	-0.171066496155	0.0437165501479	0.160288938404	-0.0321871953347	0.143116524173	0.0148612519795	-0.0108180338835	0.131632174532	0.00450450233843	0.154762993807	2.666	61.296	36.81
9	1.23018174581	0.137033743796	-0.253520858535	1.44964594544	-0.460678065489	1.17107065374	-0.0053100945626	-0.00616077237119	0.066048389998	0.0224484730517	-0.0213845376134	0.0142491336481	-0.102891884526	0.00823774215682	0.0187553198871	0.125487243175	0.0212676669074	-0.00848018426983	0.102671845319	-0.012650034691	0.167108287439	-4.362	67.736	56.223
10	4.84240758783	-1.61125481387	-1.75620618854	4.35244950141	1.23336768362	1.47636271634	0.0468192680944	0.148128826859	-0.21008660365	-0.00174983905949	-0.211839059604	0.519589024232	-0.0937361292759	-0.449277547505	0.147191623012	0.137348427457	0.0236589464744	-0.0329471258501	0.191845217927	0.00157627663664	0.245022084345	-19.592	61.275	51.196
11	0.696995534444	-0.560146788288	0.086674278599	1.02166231041	0.0251153406612	0.463810024434	-0.0232769826837	0.00228197122619	-0.140858085205	-0.0534742616317	0.0236279719699	0.117503561412	0.0786492771533	-0.0536651057501	0.0044715619798	0.1449669775	-0.0169575060908	-0.000578658757714	0.128672195534	0.00349241889305	0.21095961329	-5.058	43.578	53.931
12	7.40478308917	7.38977195804	3.28119790404	7.48466504842	3.07634572997	1.63581240289	-0.128545263943	0.145799816066	-0.0796355189812	-0.263894947059	0.145162284349	-0.149342294161	-0.0966226915628	-0.00960120992086	-0.03034629392	0.14492177934	0.0104889878602	-0.0032089473669	0.139295387038	-0.000539316608889	0.188716719732	-25.557	48.8	-6.821
13	0.604443221678	0.201766356103	-0.42091457425	0.879938337613	-0.49610906283	1.48878978785	-0.0306007565725	0.0137365086567	-0.0173587945969	0.0154737460975	0.00893759623674	0.080991194844	0.064224814805	-0.152333233336	0.0114004021595	0.119013456233	-0.0164933215837	-0.00638974101761	0.126267789447	-0.0170972258337	0.165603382436	-39.211	29.326	8.823
14	2.42387149079	0.617529950243	0.34188439151	3.81057832424	0.460837051301	2.81673320636	0.0873941424308	-0.144083620055	0.116752283979	0.140088344858	-0.140256732945	-0.195986292141	0.0318269679263	0.121207883883	0.0147893149467	0.127393293433	-0.0114729855228	0.00674520906739	0.133371020389	0.00870624696885	0.138683710393	-30.354	27.066	19.292
15	0.505041850792	0.240290227075	-0.151775219339	1.25733376049	-0.187746007788	0.660786420551	0.00205622232431	0.0645314707355	0.0414269276989	-0.00831507312979	0.0238486149789	0.175232204795	0.0372522883574	-0.163444759322	-0.0238366056318	0.133212507863	-0.00904096343021	-0.00850685453828	0.168552204627	-0.00321405534573	0.197976661203	-40.473	34.236	1.38
16	3.67250675509	2.66996608251	1.93568442549	2.34998429756	1.34311722633	2.48762768958	0.0110091088328	-0.0995570406136	0.189133282821	-0.0265502184526	-0.00462304377411	0.243757365953	-0.0561569179807	-0.0714711096052	0.0140156688258	0.195811750476	0.0506257468321	-0.0142245299263	0.135630062883	0.00300232096034	0.284923882905	-33.199	54.781	0.715
17	7.85119734917	6.54151953933	-3.0741656707	5.44607783453	-2.54891445542	1.18154322616	-0.238329658564	0.2606152917	-0.0408788276029	-0.264184449823	0.20261881663	-0.0134616441884	0.0881258761039	-0.1131798003	-0.0106704434332	0.155197061294	-0.00112012395302	0.010566236889	0.135152521097	-0.00640151261057	0.189736654775	-13.358	38.535	-5.879
18	0.671026984743	0.74797154303	0.624758377808	1.65383492512	1.36084754798	1.74958723489	-0.0160462238525	-0.0031334930859	0.0285730771283	-0.0223948465592	0.0260349279461	-0.0785821492399	-0.0743151627765	0.0406825035134	-0.0251886537459	0.144775844538	0.00366641124793	0.000746627454251	0.116637564108	0.0264338307027	0.188188156612	-0.188	59.536	0.56
19	2.99347719878	-0.0811626798142	-0.466355281075	1.17503615674	0.0796338333327	1.765304261	-0.0305871715229	-0.136576115551	-0.0058203284543	-0.0170656076021	0.00519384419305	0.0137723670207	-0.00103489103757	0.0245985774116	0.0301774078653	0.156588800319	-0.0153852674169	-0.013051226074	0.111925308059	0.00543444745307	0.137775964154	-4.564	61.288	19.736
20	2.9761560516	1.27433984717	-1.35586917311	4.5911047752	-1.71450826378	3.08841547463	0.0971226142047	-0.148386967414	-0.254984867738	0.192752726253	-0.0759486420805	0.160459410365	0.102380894692	-0.273148679736	-0.0117465884686	0.142421141995	-0.0277822518021	-0.0246327298766	0.156389828282	0.00697705204775	0.147397611909	-9.984	59.678	19.779
21	0.851499949653	-0.147497794469	-0.334603750943	1.58364217104	0.515592850929	0.90128592401	0.0177984330211	-0.0235520522944	0.0351831471974	-0.0677343012035	-0.0199568489401	-0.007892782544	-0.0889528928945	-0.0242585995362	-0.00043106673	0.153550855148	-0.0169521699745	-0.00489084641269	0.119870620757	0.00848074686218	0.181844417046	-3.042	65.646	0.381
22	4.71984131774	2.21327303683	-3.10021793117	4.00929323275	-1.914344179	2.87300787347	-0.057119711118	-0.306499599174	-0.320606684858	-0.165284118352	-0.223065886585	-0.594682115808	0.103578686626	0.418350465577	0.245417625841	0.152983303046	-0.0072614669463	-0.0121249631829	0.162994911744	-0.00741558008947	0.269872347945	12.258	59.215	5.442
23	0.905572563647	0.416920335602	-0.133512118945	1.18282177354	0.0777287424219	0.837914077051	-0.00753629633754	0.00818388576448	-0.09596335384	0.0603811216861	-0.00227531447979	-0.136416426842	0.0778609686337	0.0945277820299	0.0167242046602	0.138117440382	0.0121053609472	-0.0023268803709	0.110347341878	0.00378332262257	0.205933009362	0.648	43.862	3.221
24	4.08402532618	3.30874357054	-4.24326306007	3.46692719426	-3.70781014777	4.79849843369	0.0877625091492	-0.193626842197	-0.0647905517531	0.0233764181185	-0.0467706097871	0.00623478783718	-0.0886540540053	0.187512819432	-0.00284526090314	0.179568746184	0.0150257995428	0.00768473910546	0.151927517998	0.0102788607019	0.230277416709	-11.54	32.188	0.877

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	( CHAIN A AND RESID 4:46 )	A	4 - 46
2	X-RAY DIFFRACTION	2	( CHAIN A AND RESID 47:161 )	A	47 - 161
3	X-RAY DIFFRACTION	3	( CHAIN A AND RESID 162:265 )	A	162 - 265
4	X-RAY DIFFRACTION	4	( CHAIN A AND RESID 266:352 )	A	266 - 352
5	X-RAY DIFFRACTION	5	( CHAIN B AND RESID 4:28 )	B	4 - 28
6	X-RAY DIFFRACTION	6	( CHAIN B AND RESID 29:46 )	B	29 - 46
7	X-RAY DIFFRACTION	7	( CHAIN B AND RESID 47:114 )	B	47 - 114
8	X-RAY DIFFRACTION	8	( CHAIN B AND RESID 115:149 )	B	115 - 149
9	X-RAY DIFFRACTION	9	( CHAIN B AND RESID 150:216 )	B	150 - 216
10	X-RAY DIFFRACTION	10	( CHAIN B AND RESID 217:237 )	B	217 - 237
11	X-RAY DIFFRACTION	11	( CHAIN B AND RESID 238:352 )	B	238 - 352
12	X-RAY DIFFRACTION	12	( CHAIN C AND RESID 4:28 )	C	4 - 28
13	X-RAY DIFFRACTION	13	( CHAIN C AND RESID 29:98 )	C	29 - 98
14	X-RAY DIFFRACTION	14	( CHAIN C AND RESID 99:161 )	C	99 - 161
15	X-RAY DIFFRACTION	15	( CHAIN C AND RESID 162:313 )	C	162 - 313
16	X-RAY DIFFRACTION	16	( CHAIN C AND RESID 314:353 )	C	314 - 353
17	X-RAY DIFFRACTION	17	( CHAIN D AND RESID 4:28 )	D	4 - 28
18	X-RAY DIFFRACTION	18	( CHAIN D AND RESID 29:70 )	D	29 - 70
19	X-RAY DIFFRACTION	19	( CHAIN D AND RESID 71:114 )	D	71 - 114
20	X-RAY DIFFRACTION	20	( CHAIN D AND RESID 115:149 )	D	115 - 149
21	X-RAY DIFFRACTION	21	( CHAIN D AND RESID 150:216 )	D	150 - 216
22	X-RAY DIFFRACTION	22	( CHAIN D AND RESID 217:237 )	D	217 - 237
23	X-RAY DIFFRACTION	23	( CHAIN D AND RESID 238:329 )	D	238 - 329
24	X-RAY DIFFRACTION	24	( CHAIN D AND RESID 330:353 )	D	330 - 353

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