PDB-6ty6: Variant W229D/F290W-2 of the last common ancestor of Gram-negativ...

ID or keywords:

Entry

Database: PDB / ID: 6ty6

Title

Variant W229D/F290W-2 of the last common ancestor of Gram-negative bacteria beta-lactamase class A (GNCA4) bound to 5(6)-nitrobenzotriazole (TS-analog)

Components

Beta lactamase (GNCA4-2)

Keywords

HYDROLASE / ANTIBIOTIC RESISTANCE / ANCESTRAL RECONSTRUCTED

Function / homology

Beta-lactamase / DD-peptidase/beta-lactamase superfamily / 3-Layer(aba) Sandwich / Alpha Beta / 6-NITROBENZOTRIAZOLE / ACETATE ION / FORMIC ACID

Function and homology information

Biological species

synthetic construct (others)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.8 Å

Authors

Gavira, J.A. / Risso, V. / Sanchez-Ruiz, J.M. / Romero-Rivera, A. / Kamerlin, S.C.L. / Ortega-Munoz, M. / Santoyo-Gonzalez, F.

Funding support

Spain,

United States, 3items

Organization	Grant number	Country
Spanish Ministry of Science, Innovation, and Universities	BIO2016-74875-P	Spain
Spanish Ministry of Economy and Competitiveness	BIO2015-66426-R	Spain
Human Frontier Science Program (HFSP)	RGP0041/2017	United States

Citation

Journal: Chem Sci / Year: 2020
Title: Enhancing ade novoenzyme activity by computationally-focused ultra-low-throughput screening.
Authors: Risso, V.A. / Romero-Rivera, A. / Gutierrez-Rus, L.I. / Ortega-Munoz, M. / Santoyo-Gonzalez, F. / Gavira, J.A. / Sanchez-Ruiz, J.M. / Kamerlin, S.C.L.

History

Deposition	Jan 15, 2020	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Jun 3, 2020	Provider: repository / Type: Initial release
Revision 1.1	Sep 16, 2020	Group: Database references / Category: citation / citation_author Item: _citation.journal_id_ISSN / _citation.journal_volume ..._citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2	Jan 24, 2024	Group: Data collection / Database references / Refinement description Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	6ty6.cif.gz	421.5 KB	Display	PDBx/mmCIF format
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Validation report

Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/ty/6ty6 ftp://data.pdbj.org/pub/pdb/validation_reports/ty/6ty6	HTTPS FTP

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Related structure data

Related structure data	6twwC 6txdC 5fqkS 5fqi S: Starting model for refinement C: citing same article (ref.)
Similar structure data	2p2g-1 2p2g-2 5g1p-2 5vmq-d 6jks-1 6d66-d 2i6u-d 6jkr-2 Search similar-shape structures by Omokage search (details) Similarity search - Function & homologyF&H Search Similarity search - HomologyF&H Search

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Links

PDB pages	PDBj / wwPDB / NCBI

Deposited unit

A: Beta lactamase (GNCA4-2)

B: Beta lactamase (GNCA4-2)

C: Beta lactamase (GNCA4-2)

hetero molecules

89 kDa, 3 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

Idetical with deposited unit
defined by author&software

Unit cell

Length a, b, c (Å)	78.384, 148.357, 246.008
Angle α, β, γ (deg.)	90.000, 90.000, 90.000
Int Tables number	23
Space group name H-M	I222
Space group name Hall	I22
Symmetry operation	#1: x,y,z #2: x,-y,-z #3: -x,y,-z #4: -x,-y,z #5: x+1/2,y+1/2,z+1/2 #6: x+1/2,-y+1/2,-z+1/2 #7: -x+1/2,y+1/2,-z+1/2 #8: -x+1/2,-y+1/2,z+1/2

Components on special symmetry positions

ID	Model	Components
1	1	A-1144- HOH

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Protein , 1 types, 3 molecules ABC

#1: Protein	Beta lactamase (GNCA4-2) Mass: 28979.668 Da / Num. of mol.: 3 / Mutation: W229D, F290W Source method: isolated from a genetically manipulated source Details: 6NT is the inhibitor analog and GOL, FMT and ACT are crystallization cocktail components. Source: (gene. exp.) synthetic construct (others) / Plasmid: pET-24b(+) Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)

#1: Protein

Beta lactamase (GNCA4-2)

Mass: 28979.668 Da / Num. of mol.: 3 / Mutation: W229D, F290W
Source method: isolated from a genetically manipulated source
Details: 6NT is the inhibitor analog and GOL, FMT and ACT are crystallization cocktail components.
Source: (gene. exp.) synthetic construct (others) / Plasmid: pET-24b(+)
Production host:

Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)

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Non-polymers , 6 types, 634 molecules

#2: Chemical	ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL Mass: 92.094 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C₃H₈O₃
#3: Chemical	ChemComp-FMT / FORMIC ACID Mass: 46.025 Da / Num. of mol.: 17 / Source method: isolated from a natural source / Formula: CH₂O₂
#4: Chemical	ChemComp-ACT / ACETATE ION Mass: 59.044 Da / Num. of mol.: 5 / Source method: isolated from a natural source / Formula: C₂H₃O₂
#5: Chemical	ChemComp-NA / SODIUM ION Mass: 22.990 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Na
#6: Chemical	ChemComp-6NT / 6-NITROBENZOTRIAZOLE Mass: 164.122 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C₆H₄N₄O₂ / Feature type: SUBJECT OF INVESTIGATION
#7: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 603 / 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: 3.94 Å³/Da / Density % sol: 68.8 %
Crystal grow	Temperature: 293 K / Method: counter-diffusion / pH: 4 / Details: 5.0 M Sodium formate, 100 mM AcNa pH 4-0 / PH range: 4.0-7.0

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: ALBA / Beamline: XALOC / Wavelength: 0.97926 Å
Detector	Type: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Jun 14, 2019
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 0.97926 Å / Relative weight: 1
Reflection	Resolution: 1.8→48.48 Å / Num. obs: 130796 / % possible obs: 98.56 % / Redundancy: 5.2 % / B_iso Wilson estimate: 30.31 Å² / CC_1/2: 0.997 / CC star: 0.999 / R_merge(I) obs: 0.089 / R_pim(I) all: 0.045 / R_rim(I) all: 0.1 / Net I/σ(I): 10.77
Reflection shell	Resolution: 1.8→1.86 Å / Redundancy: 5.4 % / R_merge(I) obs: 1.44 / Mean I/σ(I) obs: 1.2 / Num. unique obs: 13114 / CC_1/2: 0.68 / CC star: 0.899 / R_pim(I) all: 0.687 / R_rim(I) all: 1.6 / % possible all: 99.39

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Processing

Software

Name	Version	Classification
EDNA		data collection
XDS		data reduction
Aimless		data scaling
PHASER		phasing
PHENIX	1.15.2_3472	refinement
PHASER		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 5FQK

5fqk

Resolution: 1.8→48.48 Å / SU ML: 0.2609 / Cross valid method: FREE R-VALUE / σ(F): 1.33 / Phase error: 27.9537
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2

	R_factor	Num. reflection	% reflection
R_free	0.2283	6624	5.07 %
R_work	0.2006	124002	-
obs	0.202	130626	98.56 %

Solvent computation

Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL

Displacement parameters

B_iso mean: 40.03 Å²

Refinement step

Cycle: LAST / Resolution: 1.8→48.48 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	6019	0	138	604	6761

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.0186	6816
X-RAY DIFFRACTION	f_angle_d	1.4386	9295
X-RAY DIFFRACTION	f_chiral_restr	0.1015	1038
X-RAY DIFFRACTION	f_plane_restr	0.0103	1259
X-RAY DIFFRACTION	f_dihedral_angle_d	9.4267	5288

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 (%)
1.8-1.82	0.4537	221	0.4209	4141	X-RAY DIFFRACTION	99.23
1.82-1.84	0.3976	230	0.382	4105	X-RAY DIFFRACTION	99.45
1.84-1.86	0.3608	219	0.3286	4144	X-RAY DIFFRACTION	99.5
1.86-1.89	0.322	236	0.304	4095	X-RAY DIFFRACTION	99.7
1.89-1.91	0.2796	226	0.2735	4172	X-RAY DIFFRACTION	99.73
1.91-1.94	0.2815	218	0.2613	4093	X-RAY DIFFRACTION	99.75
1.94-1.97	0.2897	229	0.2564	4151	X-RAY DIFFRACTION	99.61
1.97-2	0.2996	207	0.2457	4186	X-RAY DIFFRACTION	99.82
2-2.03	0.262	201	0.2385	4124	X-RAY DIFFRACTION	99.47
2.03-2.06	0.2628	227	0.2386	4210	X-RAY DIFFRACTION	99.66
2.06-2.1	0.2428	220	0.243	4118	X-RAY DIFFRACTION	99.77
2.1-2.13	0.259	213	0.2414	4153	X-RAY DIFFRACTION	99.41
2.13-2.18	0.2863	231	0.2392	4131	X-RAY DIFFRACTION	99.48
2.18-2.22	0.2718	204	0.2244	4141	X-RAY DIFFRACTION	99.29
2.22-2.27	0.2808	226	0.2264	4155	X-RAY DIFFRACTION	99.61
2.27-2.32	0.2589	231	0.22	4173	X-RAY DIFFRACTION	99.41
2.32-2.38	0.2611	232	0.2259	4129	X-RAY DIFFRACTION	99.2
2.38-2.44	0.2758	241	0.2217	4143	X-RAY DIFFRACTION	99.25
2.44-2.51	0.236	217	0.2256	4123	X-RAY DIFFRACTION	99.34
2.51-2.6	0.2595	219	0.217	4144	X-RAY DIFFRACTION	99.05
2.6-2.69	0.2368	217	0.2243	4117	X-RAY DIFFRACTION	98.1
2.69-2.8	0.2516	211	0.2271	4131	X-RAY DIFFRACTION	97.93
2.8-2.92	0.272	228	0.2305	4059	X-RAY DIFFRACTION	96.95
2.92-3.08	0.2734	223	0.2203	4048	X-RAY DIFFRACTION	96.85
3.08-3.27	0.2291	208	0.2037	4094	X-RAY DIFFRACTION	96.31
3.27-3.52	0.2052	221	0.1794	4055	X-RAY DIFFRACTION	96.37
3.52-3.88	0.171	196	0.1564	4041	X-RAY DIFFRACTION	94.98
3.88-4.44	0.1686	214	0.1431	4060	X-RAY DIFFRACTION	95.02
4.44-5.59	0.1586	231	0.1425	4196	X-RAY DIFFRACTION	97.43
5.59-48.48	0.201	227	0.1778	4370	X-RAY DIFFRACTION	97.79

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	1.32062591543	-0.135248533669	0.112194433394	2.72679544943	0.459868721151	2.27142230196	0.0527380068774	0.0890261494166	-0.127426454321	-0.490071026301	-0.00363712768733	-0.0369189658187	0.547764327127	0.0047255039053	-0.011760642492	0.394532923508	-0.0416041530337	-0.000534283869238	0.249355699133	-0.00712452499365	0.277973020663	21.4147929488	-5.2587607366	38.7006477472
2	0.797165932791	-0.109767308316	-0.196832404777	2.0485625997	0.627629687244	1.72439345644	-0.0230628042244	-0.0557539763051	0.0216302440169	-0.0582497988258	-0.00295467681663	0.142178620143	-0.00299273875201	-0.141832404416	-0.0017128947285	0.1977073587	0.00305723868269	0.0137919074424	0.25292345105	0.0124058337319	0.272811143883	18.9691955286	17.7004820427	51.8267393697
3	0.46867847696	-0.262247074667	0.270762868904	3.10288988393	0.435715842969	1.90198388257	-0.0984702417913	-0.0313817995351	-0.0836185769173	0.169682145049	0.159433392578	-0.0141703001175	0.25059334326	-0.0336289640677	-0.0130422187707	0.18731522883	0.0303638179419	0.0207596752846	0.254999173339	0.0298910663873	0.252447830955	24.0410429194	4.03766088789	57.1417888385
4	1.7743822277	-0.193084620381	0.417234879279	2.49576870575	-0.316816795609	2.74594097299	-0.195435187676	-0.0685038380633	-0.0731531547729	-0.0942814208431	0.0874799410172	0.152578852389	0.0508808975374	-0.263346607437	0.0106337988716	0.252743027762	-0.0383885751416	0.0268416216095	0.238866617051	0.0278609180742	0.308458881319	19.8848289369	-0.773883766755	47.4559381583
5	1.66413709492	0.762360368189	0.110487051714	4.04088934722	-0.207962747626	4.10925245259	0.00682719893601	0.219449628659	-0.506046534025	-0.291970424437	-0.0948740817157	0.0150136289148	0.450250539102	0.137428180801	0.0227939979353	0.314521848466	-0.0813628553382	0.0283132429905	0.210936440478	0.0324936714076	0.273936038139	20.4436808504	-10.5014110814	47.4318633121
6	0.591201169366	0.255952776691	0.341639843721	1.08809486662	-0.131185663545	1.42227352982	0.0330170713835	-0.290674443907	0.0367111471966	0.167138509724	-0.0513270836852	0.147699366555	-0.108734992048	-0.305305969722	-0.0179438907374	0.246222314836	0.0813630815077	0.0365104777264	0.347527572947	-0.0226176042879	0.293197394826	12.8856290473	43.6256313732	66.7832172098
7	1.77318748699	0.0878480404361	-0.767057376948	0.962257022203	0.279598542538	0.975996259807	0.0143528150825	0.048987580108	0.0834197162309	-0.365108684882	0.0791896530802	-0.238096144238	-0.404172486446	0.22208909714	0.0335375076046	0.4352993431	0.000591692055574	0.0817398684412	0.326872400533	-0.0105477650695	0.340727198571	29.241356452	45.1359262245	39.7339773775
8	0.16704016981	0.0523935949024	-0.0985528827074	0.421576822626	-0.00922728969361	0.378811225809	0.117498195939	0.277978689512	0.147898141473	-0.574506973183	-0.0832329561522	0.260936278964	-0.726910372778	-0.396055385541	-0.387196315207	0.673028757538	0.103218741663	0.0278800297165	0.415392426907	0.0441432002503	0.362516941305	16.8653280436	52.4021362006	31.6785711766
9	0.842358167926	-0.261378706576	-0.515533261474	1.35193534471	0.430547254619	1.10521356423	0.0402837743311	-0.0172931771477	0.12663135388	-0.245774672419	0.0556949028597	-0.0899666573676	-0.239219724017	-0.0542637467728	-0.0392457558365	0.258588573521	0.0280481054192	0.0221513320217	0.259402416486	-0.0094842229171	0.256032194541	21.8565093562	46.4398328773	51.0161204707
10	0.094626735059	-0.0742980419555	-0.13592315861	0.293526914844	0.00487661375262	0.308430325729	-0.203183337421	0.337919613213	0.204584614505	-0.164819087444	0.083211290482	0.237522093615	-0.152886873017	-0.247856477039	-0.0733300491547	0.239925976127	0.0880390532121	-0.00930121590708	0.420430456139	-0.0224214213153	0.380641539976	8.95524290717	46.9358404917	58.4297395375
11	3.70941568791	0.529085334409	2.18295437354	4.07620262704	0.210874191751	2.55742344077	0.158668201009	-0.147059815955	0.470605214955	0.183016651551	-0.219521371042	-0.136674388594	0.145674012665	-0.0806850878579	0.0163681982405	0.291945676435	0.0822191858937	0.0580709868462	0.368372398344	-0.0499812823416	0.325729922011	13.8788130774	53.8667270431	67.6771722674
12	2.73632221265	-0.0293238428367	-1.68894471075	2.97534525189	-0.822870816836	3.89573671411	0.0375463684548	0.436088743097	0.445700152399	-0.288463773923	0.189668825604	-0.0965896163785	-0.370381752851	-0.035257833721	0.431279387583	0.927202337308	0.139748088739	0.115898408141	0.437239137041	0.123354455537	0.348291054314	25.8794978751	48.3392546742	5.41926464841
13	0.238195703555	0.359559282538	-0.400869549378	1.52733425968	-0.555015124915	0.853073743143	0.115837106555	-0.0258473418364	0.0165329693712	-0.121583102233	0.048104671119	-0.283576181465	-0.613312727697	0.212479622294	0.166206156139	0.666043489039	0.0119149514136	0.138216514595	0.439034653865	-0.00355897731749	0.297174260369	32.5018980976	43.2693361981	17.0460522226
14	0.380753861807	-0.142706732404	-0.37070121311	1.18887743668	-0.302305430452	0.511114333857	0.0237279903429	0.065835736753	-0.164193707449	-0.454932134136	-0.0288177773589	0.0630810141326	0.203829650241	-0.181810255292	-0.0540516221283	0.527817144677	-0.0110617905937	0.0648314919037	0.326614118603	-0.0289871880255	0.321351681706	28.0797459211	14.2345604722	25.0483916305
15	0.304868995365	0.0551440615406	-0.227654942536	0.647227870365	-0.0442881141294	0.357015662265	0.0337710606311	0.102514020527	-0.00879777029903	-0.441169796265	-0.0125161415865	0.0276035047049	-0.0288447550028	-0.181456040788	-0.0275205199454	0.472593132254	0.0111137246682	0.0528384644351	0.354151947644	0.000213808455329	0.289453375842	25.25181224	25.7204056965	26.3879441211
16	0.33409564268	-0.235664327927	-0.359095570011	0.553876206535	-0.244321906795	0.819070546908	0.0687244918786	0.0432616984594	-0.0347840293544	-0.456334769358	-0.0164256616426	-0.187084645979	-0.0728787888165	0.21954470071	-0.00881921239621	0.60649365225	-0.00775788402387	0.126537640803	0.377265877487	-0.0152575326529	0.327359378276	37.2002098439	28.7851124161	16.4069802143
17	2.31278449868	0.338326289433	-1.60603878481	0.878886311151	-0.341009887327	1.95105057307	-0.154644321383	0.454888238216	0.0240706502034	-0.280624435464	0.154946632709	-0.0786384536682	-0.2222209554	0.0594476470855	0.0597454846283	0.726600713825	0.0036972266745	0.12084829896	0.4466716123	0.0110296598051	0.347419921054	35.1337733503	33.3070055976	12.3743129082
18	2.3309190543	-0.645608445036	0.742981820779	0.293962940272	-0.675848522098	2.80823921972	0.181454012987	0.164922971651	-0.348782994814	-0.477578519891	0.0292961508333	0.184259864406	-0.326031972343	-0.383379150426	0.417261715895	0.841557035096	0.107890539811	0.165004818009	0.30057658207	0.0254524508127	0.148848075607	29.9661651564	38.8699798039	7.51170573707

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	chain 'A' and (resid 27 through 68 )
2	X-RAY DIFFRACTION	2	chain 'A' and (resid 69 through 195 )
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 196 through 238 )
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 240 through 271 )
5	X-RAY DIFFRACTION	5	chain 'A' and (resid 272 through 294 )
6	X-RAY DIFFRACTION	6	chain 'B' and (resid 27 through 68 )
7	X-RAY DIFFRACTION	7	chain 'B' and (resid 69 through 98 )
8	X-RAY DIFFRACTION	8	chain 'B' and (resid 99 through 118 )
9	X-RAY DIFFRACTION	9	chain 'B' and (resid 119 through 258 )
10	X-RAY DIFFRACTION	10	chain 'B' and (resid 259 through 271 )
11	X-RAY DIFFRACTION	11	chain 'B' and (resid 272 through 295 )
12	X-RAY DIFFRACTION	12	chain 'C' and (resid 27 through 40 )
13	X-RAY DIFFRACTION	13	chain 'C' and (resid 41 through 68 )
14	X-RAY DIFFRACTION	14	chain 'C' and (resid 69 through 118 )
15	X-RAY DIFFRACTION	15	chain 'C' and (resid 119 through 179 )
16	X-RAY DIFFRACTION	16	chain 'C' and (resid 180 through 238 )
17	X-RAY DIFFRACTION	17	chain 'C' and (resid 240 through 258 )
18	X-RAY DIFFRACTION	18	chain 'C' and (resid 259 through 294 )

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