PDB-7k4q: Crystal structure of Kemp Eliminase HG3 in complex with the trans...

ID or keywords:

Entry

Database: PDB / ID: 7k4q

Title

Crystal structure of Kemp Eliminase HG3 in complex with the transition state analog 6-nitrobenzotriazole

Components

Endo-1,4-beta-xylanase

Xylanase

Keywords

HYDROLASE /

kemp elimination /

directed evolution

Function / homology

Function and homology information

endo-1,4-beta-xylanase activity /

endo-1,4-beta-xylanase / xylan catabolic process
Similarity search - Function

Biological species

Thermoascus aurantiacus (fungus)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.28 Å

Authors

Padua, R.A.P. / Otten, R. / Bunzel, A. / Nguyen, V. / Pitsawong, W. / Patterson, M. / Sui, S. / Perry, S.L. / Cohen, A.E. / Hilvert, D. / Kern, D.

Funding support

United States,

Switzerland, 11items

Organization	Grant number	Country
Department of Energy (DOE, United States)	DE-FG02-05ER15699	United States
Damon Runyon Cancer Research Foundation	DRG-2114-12	United States
Department of Energy (DOE, United States)	DE-AC02-76SF00515	United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)	P41GM103393	United States
Howard Hughes Medical Institute (HHMI)		United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)	P30 GM124169	United States
Department of Energy (DOE, United States)	DE-AC02-05CH11231	United States
National Institutes of Health/Office of the Director	S10OD021832	United States
Swiss National Science Foundation		Switzerland
Other government		Switzerland
National Science Foundation (NSF, United States)	BioXFEL STC (1231306)	United States

Citation

Journal: Science / Year: 2020
Title: How directed evolution reshapes the energy landscape in an enzyme to boost catalysis.
Authors: Otten, R. / Padua, R.A.P. / Bunzel, H.A. / Nguyen, V. / Pitsawong, W. / Patterson, M. / Sui, S. / Perry, S.L. / Cohen, A.E. / Hilvert, D. / Kern, D.

History

Deposition	Sep 16, 2020	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Dec 2, 2020	Provider: repository / Type: Initial release
Revision 1.1	Dec 30, 2020	Group: Database references / Category: citation / citation_author Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.2	Oct 18, 2023	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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Download

PDBx/mmCIF format	7k4q.cif.gz	444.7 KB	Display	PDBx/mmCIF format
PDB format	pdb7k4q.ent.gz	303.4 KB	Display	PDB format
PDBx/mmJSON format	7k4q.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/k4/7k4q ftp://data.pdbj.org/pub/pdb/validation_reports/k4/7k4q	HTTPS FTP

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Related structure data

Related structure data	7k4pC 7k4rC 7k4sC 7k4tC 7k4uC 7k4vC 7k4wC 7k4xC 7k4yC 7k4zC 4bs0S S: Starting model for refinement C: citing same article (ref.)
Similar structure data	7k4p-1 5rgb-2 7k4x-1 7k4s-d 5h2u-3 5rg9-2 6tu6-9 7k4x-2 1b3w-d 6ss1-1 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
Related items in Molecule of the Month	#217 - Jan 2018 Opioid Receptors similarity (1)

Deposited unit

A: Endo-1,4-beta-xylanase

B: Endo-1,4-beta-xylanase

hetero molecules

66.9 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: Endo-1,4-beta-xylanase

hetero molecules

defined by author&software
Evidence: gel filtration
33.5 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

B: Endo-1,4-beta-xylanase

hetero molecules

defined by author&software
33.5 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

Unit cell

Length a, b, c (Å)	50.640, 58.970, 88.760
Angle α, β, γ (deg.)	90.000, 104.940, 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

#1: Protein	Endo-1,4-beta-xylanase / Xylanase / Xylanase / 1 / 4-beta-D-xylan xylanohydrolase / TAXI Mass: 33200.352 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Thermoascus aurantiacus (fungus) / Gene: XYNA / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P23360, endo-1,4-beta-xylanase
#2: Chemical	ChemComp-6NT / 6-NITROBENZOTRIAZOLE Mass: 164.122 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₆H₄N₄O₂
#3: Chemical	ChemComp-SO4 / SULFATE ION / Sulfate Mass: 96.063 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: SO₄
#4: Water	ChemComp-HOH / water / Water Mass: 18.015 Da / Num. of mol.: 781 / 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: 1.93 Å³/Da / Density % sol: 36.22 %
Crystal grow	Temperature: 291 K / Method: vapor diffusion, sitting drop / Details: 2 M ammonium sulfate and 5% isopropanol

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: SSRL / Beamline: BL14-1 / Wavelength: 0.9537 Å
Detector	Type: MARMOSAIC 325 mm CCD / Detector: CCD / Date: Feb 17, 2019
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 0.9537 Å / Relative weight: 1
Reflection	Resolution: 1.277→48.59 Å / Num. obs: 126111 / % possible obs: 96.27 % / Redundancy: 6.4 % / B_iso Wilson estimate: 11.43 Å² / CC_1/2: 0.999 / Net I/σ(I): 10.07
Reflection shell	Resolution: 1.277→1.323 Å / Num. unique obs: 12643 / CC_1/2: 0.919

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Processing

Software

Name	Version	Classification
PHENIX	dev_3965	refinement
Coot		model building
iMOSFLM		data reduction
Aimless		data scaling
PHASER		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 4bs0

4bs0

Resolution: 1.28→48.59 Å / SU ML: 0.1233 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 23.8699
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2

	R_factor	Num. reflection	% reflection
R_free	0.19	3930	1.59 %
R_work	0.1681	243920	-
obs	0.1685	125995	96.22 %

Solvent computation

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

Displacement parameters

B_iso mean: 15.51 Å²

Refinement step

Cycle: LAST / Resolution: 1.28→48.59 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4543	0	34	781	5358

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.0046	5032
X-RAY DIFFRACTION	f_angle_d	0.8206	6920
X-RAY DIFFRACTION	f_chiral_restr	0.0661	770
X-RAY DIFFRACTION	f_plane_restr	0.0067	924
X-RAY DIFFRACTION	f_dihedral_angle_d	5.9596	708

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 (%)
1.28-1.29	0.3003	141	0.2871	8753	X-RAY DIFFRACTION	97.14
1.29-1.31	0.301	143	0.2802	8803	X-RAY DIFFRACTION	97.03
1.31-1.33	0.2899	147	0.259	8824	X-RAY DIFFRACTION	97.34
1.33-1.34	0.261	139	0.2581	8725	X-RAY DIFFRACTION	96
1.34-1.36	0.2704	146	0.2487	8752	X-RAY DIFFRACTION	96.13
1.36-1.38	0.2274	141	0.2422	8562	X-RAY DIFFRACTION	94.62
1.38-1.41	0.2581	141	0.2363	8481	X-RAY DIFFRACTION	94.17
1.41-1.43	0.2175	123	0.2292	8232	X-RAY DIFFRACTION	91.66
1.43-1.45	0.2063	141	0.2192	8308	X-RAY DIFFRACTION	91.11
1.45-1.48	0.2391	139	0.2013	8834	X-RAY DIFFRACTION	97.35
1.48-1.51	0.2295	135	0.1983	8796	X-RAY DIFFRACTION	97.52
1.51-1.54	0.1936	147	0.1871	8873	X-RAY DIFFRACTION	98.21
1.54-1.57	0.1768	149	0.1833	8742	X-RAY DIFFRACTION	96.89
1.57-1.61	0.2085	136	0.1789	8836	X-RAY DIFFRACTION	97.25
1.61-1.65	0.185	142	0.1697	8757	X-RAY DIFFRACTION	96.67
1.65-1.69	0.2045	144	0.1701	8714	X-RAY DIFFRACTION	96.04
1.69-1.74	0.1744	146	0.168	8672	X-RAY DIFFRACTION	95.73
1.74-1.8	0.1582	136	0.1584	8606	X-RAY DIFFRACTION	95.16
1.8-1.86	0.1919	131	0.1641	8288	X-RAY DIFFRACTION	91.96
1.86-1.94	0.2311	133	0.1716	8369	X-RAY DIFFRACTION	92.2
1.94-2.03	0.1825	149	0.1556	9008	X-RAY DIFFRACTION	99.62
2.03-2.13	0.1804	145	0.1491	9015	X-RAY DIFFRACTION	99.66
2.13-2.27	0.1676	143	0.1511	8986	X-RAY DIFFRACTION	99.05
2.27-2.44	0.1679	147	0.1438	8944	X-RAY DIFFRACTION	99.15
2.44-2.69	0.1645	136	0.1462	8965	X-RAY DIFFRACTION	98.53
2.69-3.08	0.233	140	0.1514	8624	X-RAY DIFFRACTION	95.47
3.08-3.88	0.1533	131	0.1432	8485	X-RAY DIFFRACTION	93.37
3.88-48.59	0.1563	139	0.15	8966	X-RAY DIFFRACTION	99.18

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	2.59635621237	2.02771475835	-0.268418472966	6.59811776529	0.646226477062	2.61858048484	0.0472030589433	-0.151289453342	-0.0352691332648	0.163299128095	-0.128545197223	0.222063467703	0.118505718538	-0.252585348107	0.074553499902	0.0647959674074	-0.0125002279808	-0.0113094621168	0.107629462953	-0.00829223263554	0.0541688550879	-20.0980161921	22.3136271779	9.13072527932
2	2.44893885455	0.680046625338	-1.2993205536	0.449969240497	-0.229998021456	1.73124133104	0.108461955034	-0.203985181705	0.103202515635	0.0519972036861	-0.0785356243457	0.0167173232704	-0.0559710295211	0.0569685883699	-0.0328496423926	0.0833596115242	-0.00717280375088	-0.0137124645948	0.0891064137055	-0.00744812125768	0.0838478385138	-0.488146435409	32.0605063338	7.98639536191
3	3.12531053803	1.65996707563	-1.48317631823	1.87443751462	-1.79020147028	3.2206554985	-0.0213282385822	-0.131326966193	-0.0838299872446	-0.0076632804012	-0.0823361378753	-0.10167904484	0.0606169335952	0.18477303152	0.124834635845	0.0683694408381	0.0127430170847	-0.0113612162123	0.0510867373224	-0.010322064661	0.069621879957	6.43487543437	24.087920559	1.20725833499
4	1.35160698327	0.212079684766	-0.102379448639	0.979398310855	-0.309309724806	0.970003961059	0.000446068467183	0.126560345223	0.0252860528631	-0.0570752872809	-0.0245604464499	-0.0220862163252	-0.0253040842974	0.0237484427349	0.0314194764015	0.0770089624845	0.0021558679738	-0.00730226618974	0.0736414057622	-0.00703395342519	0.0678947667491	-1.5063074788	27.0267919288	-9.81752854031
5	7.64647506099	-6.2492660088	0.597459626703	7.4267811654	-0.98248653569	0.789553723827	0.12791192763	0.335509503907	-0.21904873032	-0.320291976047	-0.0723788647704	0.170222937923	0.0892812620911	-0.043971145397	-0.0489350246211	0.121504693443	0.0124728675659	-0.00602073808971	0.113880021218	-0.0150920086114	0.0663231907445	-6.87232510379	21.9677701824	-16.1794944449
6	1.72495725427	-0.843478697345	-0.453436188003	1.4188997707	0.424892527592	0.995559632143	0.0210915559301	0.0292278011812	0.149501902899	-0.0157142197083	-0.0168133215713	-0.0271534301724	-0.0317422211649	-0.0760170164986	-0.0232801760062	0.0692125396706	-0.00304823427597	-0.0124101409072	0.0764225061931	-0.00222189093816	0.0741992950662	-17.2849348635	28.243190855	-2.95506643947
7	3.21657243738	-0.516234053976	-0.0375767561904	2.20483153951	0.605885152429	4.44567264762	0.0595670201389	-0.410403621575	0.332838282622	0.221179111538	-0.0537308212883	0.02111921898	-0.189619123017	-0.149886955039	-0.00659330447893	0.0826437015561	0.0274948268481	-0.00835596801824	0.185310639042	-0.0683329930597	0.123855631469	-24.8801591927	34.3430299971	9.08372339287
8	2.84190326953	1.47274928136	0.442575458454	3.77776856131	1.1331670706	2.75215684231	0.00927445435811	-0.243419327512	0.191159264259	0.13637521316	-0.0617412262364	0.121179952385	-0.00915891365018	-0.0595722691717	0.0533521873272	0.0766838929103	0.0181080681593	-0.00972388499679	0.0666082859898	-0.00464049678375	0.0688149095815	-1.74490863872	30.0773682225	55.1336842724
9	1.26845996437	0.516864181931	-1.06369508484	0.399452012539	-0.597368250278	1.67740548593	0.0399108632667	-0.122336824625	0.0463959495463	0.0315061072531	-0.0305524402917	-0.0332910089338	-0.057710685728	0.121907018154	-0.0408116429309	0.0909545994703	6.20655793683E-5	-0.0155436644049	0.0769104207423	-0.00515029211882	0.0978837041595	17.1124065649	34.2714495821	48.6446062247
10	1.85616100993	0.743678401601	-1.06515447802	0.70932750405	-0.733733564726	1.96758871292	-0.0139607658305	-0.0726749527396	-0.0986648522829	0.00712982602134	-0.0622648670803	-0.0534385036167	0.0730906729158	0.147486916465	0.133004609261	0.0919164105685	0.0109355779736	-0.0129388142133	0.0711931893818	-0.0063387657336	0.113785781099	20.2000739891	26.5795293852	43.8837697271
11	1.44689797097	0.140634940043	0.229903951733	0.433692878479	-0.219903872985	0.692711020984	-0.0301675291494	0.122915087423	0.0267184480545	-0.0540838976827	0.0274404992459	-0.0278335478264	0.0149776773815	0.0595282762684	0.00964333162218	0.104428634851	-0.0095301131237	0.000398408590247	0.0771382095953	-0.0122471645906	0.0956188336208	12.4423348685	29.5041925554	32.9076031889
12	7.88752263466	-7.31036678269	2.04416625455	8.33830645753	-2.14954548972	1.4231144475	0.130939689727	0.360006720552	-0.197757421214	-0.214182921729	-0.0736761325121	0.0911850047471	0.112425311546	0.0950190058684	-0.0426659329367	0.108876435009	-0.018927800951	-0.00189491752523	0.0913846253305	-0.0174974594137	0.0691213413268	6.13563491058	25.0828430211	26.1663457841
13	1.69657035146	-0.84470517024	-0.814458363014	1.00492710253	0.744360160229	1.15370454218	0.0138313553757	0.0970667988862	0.063307305342	-0.0127228355392	-0.00146893122869	-0.0149880059515	0.0166312145116	-0.087874483797	-0.0196073948543	0.0939907126059	-0.0120028031647	-0.0144140950673	0.064245030284	0.0110945070516	0.0869028036549	-3.33072345367	29.7478537806	39.2687037272
14	5.07702999548	0.648367066399	0.531267267406	3.82673779744	1.19367145075	4.52409774602	0.0289845068576	-0.183700223748	0.367535051178	0.044668704309	-0.0242616925039	-0.165467691163	-0.298029790188	-0.0596996800335	-0.00667884811318	0.0757986904358	0.0381688324454	-0.0051626992387	0.106611912964	-0.0204465174902	0.11829608919	-9.32952993021	38.5300293712	51.3598436463

Refinement TLS group

Refine-ID: X-RAY DIFFRACTION

ID	Refine TLS-ID	Selection details	Auth asym-ID	Label asym-ID	Auth seq-ID	Label seq-ID
1	1	chain 'A' and (resid 4 through 22 )	A	A	4 - 22	1 - 19
2	2	chain 'A' and (resid 23 through 100 )	A	A	23 - 100	20 - 97
3	3	chain 'A' and (resid 101 through 130 )	A	A	101 - 130	98 - 127
4	4	chain 'A' and (resid 131 through 181 )	A	A	131 - 181	128 - 178
5	5	chain 'A' and (resid 182 through 197 )	A	A	182 - 197	179 - 194
6	6	chain 'A' and (resid 198 through 273 )	A	A	198 - 273	195 - 270
7	7	chain 'A' and (resid 274 through 303 )	A	A	274 - 303	271 - 297
8	8	chain 'B' and (resid 4 through 45 )	B	C	4 - 45	1 - 42
9	9	chain 'B' and (resid 46 through 100 )	B	C	46 - 100	43 - 97
10	10	chain 'B' and (resid 101 through 130 )	B	C	101 - 130	98 - 127
11	11	chain 'B' and (resid 131 through 181 )	B	C	131 - 181	128 - 178
12	12	chain 'B' and (resid 182 through 196 )	B	C	182 - 196	179 - 193
13	13	chain 'B' and (resid 197 through 270 )	B	C	197 - 270	194 - 267
14	14	chain 'B' and (resid 271 through 303 )	B	C	271 - 303	268 - 300

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