PDB-8usg: Crystal Structure of Kemp Eliminase HG630 in unbound state, 280 K

ID or keywords:

Entry

Database: PDB / ID: 8usg

Title

Crystal Structure of Kemp Eliminase HG630 in unbound state, 280 K

Components

Kemp eliminase

Keywords

HYDROLASE / De novo enzyme design / computational protein design / Kemp eliminases / biocatalysis / directed evolution / X-ray crystallography

Biological species

Thermoascus aurantiacus (fungus)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.37 Å

Authors

Seifinoferest, B.

Funding support

Canada,

France,

United States, 4items

Organization	Grant number	Country
Natural Sciences and Engineering Research Council (NSERC, Canada)		Canada
Canada Foundation for Innovation		Canada
Human Frontier Science Program (HFSP)		France
Department of Energy (DOE, United States)		United States

Citation

Journal: J.Am.Chem.Soc. / Year: 2024
Title: Design of Efficient Artificial Enzymes Using Crystallographically Enhanced Conformational Sampling.
Authors: Rakotoharisoa, R.V. / Seifinoferest, B. / Zarifi, N. / Miller, J.D.M. / Rodriguez, J.M. / Thompson, M.C. / Chica, R.A.

History

Deposition	Oct 27, 2023	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Dec 6, 2023	Provider: repository / Type: Initial release
Revision 1.1	Jun 19, 2024	Group: Database references / Category: citation / citation_author Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _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 / _citation.year / _citation_author.identifier_ORCID
Revision 1.2	Nov 6, 2024	Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature / Item: _pdbx_entry_details.has_protein_modification

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	8usg.cif.gz	462.5 KB	Display	PDBx/mmCIF format
PDB format	pdb8usg.ent.gz	316.8 KB	Display	PDB format
PDBx/mmJSON format	8usg.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	8usg_validation.pdf.gz	437.2 KB	Display	wwPDB validaton report
Full document	8usg_full_validation.pdf.gz	439 KB	Display
Data in XML	8usg_validation.xml.gz	28.6 KB	Display
Data in CIF	8usg_validation.cif.gz	44.7 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/us/8usg ftp://data.pdbj.org/pub/pdb/validation_reports/us/8usg	HTTPS FTP

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Related structure data

Related structure data	8useC 8usfC 8ushC 8usiC 8usjC 8uskC 8uslC C: citing same article (ref.)

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Links

PDB pages	PDBj / wwPDB / NCBI

Deposited unit

A: Kemp eliminase

B: Kemp eliminase

hetero molecules

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

1

A: Kemp eliminase

hetero molecules

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

2

B: Kemp eliminase

hetero molecules

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

Unit cell

Length a, b, c (Å)	51.116, 77.521, 93.872
Angle α, β, γ (deg.)	90.000, 105.444, 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	Kemp eliminase Mass: 34252.406 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Thermoascus aurantiacus (fungus) / Production host: Escherichia coli (E. coli)
#2: Chemical	ChemComp-SO4 / SULFATE ION Mass: 96.063 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: SO₄
#3: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 557 / Source method: isolated from a natural source / Formula: H₂O
Has ligand of interest	N
Has protein modification	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: 2.62 Å³/Da / Density % sol: 53 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 4.8 Details: 10.6 mg/mL protein, 100 mM sodium acetate, pH 4.8, 1.4 M ammonium sulfate

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Data collection

Diffraction	Mean temperature: 280 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: ALS / Beamline: 8.3.1 / Wavelength: 1.1 Å
Detector	Type: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Mar 17, 2022
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 1.1 Å / Relative weight: 1
Reflection	Resolution: 1.32→90.6 Å / Num. obs: 164672 / % possible obs: 99.7 % / Redundancy: 3.3 % / B_iso Wilson estimate: 13.06 Å² / CC_1/2: 0.995 / R_merge(I) obs: 0.08 / R_pim(I) all: 0.052 / R_rim(I) all: 0.096 / Net I/σ(I): 9.4
Reflection shell	Resolution: 1.32→1.34 Å / Redundancy: 3.2 % / R_merge(I) obs: 0.897 / Mean I/σ(I) obs: 1.1 / Num. unique obs: 8177 / CC_1/2: 0.477 / R_pim(I) all: 0.585 / % possible all: 98.9

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Processing

Software

Name	Version	Classification
xia2	0.5.492	data reduction
DIALS	Xia2 0.5.492	data reduction
Aimless	Xia2 0.5.492	data scaling
Coot	0.8.9.236	model building
PHASER	v1.13.2998	phasing
PHENIX	v1.13-2998	refinement

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 1.37→45.24 Å / SU ML: 0.128 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 13.4204
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2

	R_factor	Num. reflection	% reflection
R_free	0.1408	7216	4.92 %
R_work	0.1274	139565	-
obs	0.1281	146781	99.2 %

Solvent computation

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

Displacement parameters

B_iso mean: 21.16 Å²

Refinement step

Cycle: LAST / Resolution: 1.37→45.24 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	4570	0	10	557	5137

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.0067	5330
X-RAY DIFFRACTION	f_angle_d	1.0029	7376
X-RAY DIFFRACTION	f_chiral_restr	0.0747	819
X-RAY DIFFRACTION	f_plane_restr	0.0067	1011
X-RAY DIFFRACTION	f_dihedral_angle_d	12.0566	1960

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 (%)
1.37-1.39	0.2824	255	0.2725	4243	X-RAY DIFFRACTION	90.25
1.39-1.4	0.257	201	0.2487	4511	X-RAY DIFFRACTION	97.07
1.4-1.42	0.2379	238	0.2232	4669	X-RAY DIFFRACTION	99.05
1.42-1.44	0.2307	244	0.2143	4584	X-RAY DIFFRACTION	99.18
1.44-1.46	0.2215	244	0.204	4646	X-RAY DIFFRACTION	99.47
1.46-1.48	0.2016	200	0.1983	4674	X-RAY DIFFRACTION	99.41
1.48-1.5	0.2266	233	0.1864	4683	X-RAY DIFFRACTION	99.27
1.5-1.52	0.1848	273	0.1754	4579	X-RAY DIFFRACTION	99.37
1.52-1.54	0.2052	224	0.1707	4695	X-RAY DIFFRACTION	99.45
1.54-1.57	0.1754	251	0.1626	4607	X-RAY DIFFRACTION	99.82
1.57-1.6	0.1816	230	0.1591	4710	X-RAY DIFFRACTION	99.78
1.6-1.62	0.1834	264	0.1584	4634	X-RAY DIFFRACTION	99.74
1.62-1.66	0.1565	279	0.15	4622	X-RAY DIFFRACTION	99.72
1.66-1.69	0.1675	232	0.1444	4693	X-RAY DIFFRACTION	99.78
1.69-1.73	0.1711	259	0.14	4617	X-RAY DIFFRACTION	99.63
1.73-1.77	0.1582	227	0.1339	4700	X-RAY DIFFRACTION	99.62
1.77-1.81	0.1403	214	0.1274	4688	X-RAY DIFFRACTION	99.65
1.81-1.86	0.1367	239	0.1188	4651	X-RAY DIFFRACTION	99.67
1.86-1.91	0.1419	216	0.1141	4705	X-RAY DIFFRACTION	99.72
1.91-1.98	0.1287	271	0.1124	4665	X-RAY DIFFRACTION	99.68
1.98-2.05	0.1403	257	0.1161	4647	X-RAY DIFFRACTION	99.74
2.05-2.13	0.137	232	0.1117	4678	X-RAY DIFFRACTION	99.57
2.13-2.23	0.1107	266	0.1058	4643	X-RAY DIFFRACTION	99.59
2.23-2.34	0.1115	210	0.1067	4708	X-RAY DIFFRACTION	99.49
2.34-2.49	0.1245	257	0.1088	4657	X-RAY DIFFRACTION	99.66
2.49-2.68	0.1356	209	0.1209	4736	X-RAY DIFFRACTION	99.66
2.68-2.95	0.1335	223	0.1263	4707	X-RAY DIFFRACTION	99.72
2.95-3.38	0.1402	264	0.1309	4703	X-RAY DIFFRACTION	99.74
3.38-4.26	0.1117	249	0.1056	4727	X-RAY DIFFRACTION	99.76
4.26-45.24	0.1294	255	0.1188	4783	X-RAY DIFFRACTION	99.7

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	1.10940292452	0.180927144454	0.560146620398	1.3146489508	0.510441449902	2.3940234638	-0.0361693613676	0.0216096596241	0.0903663672804	-0.0457026995947	-0.0148128641698	-0.0565322226797	-0.0618643859823	0.052037849351	0.0439692952829	0.174175125029	0.00154966034676	0.00446858505773	0.141502624162	0.00900452960824	0.152732673792	4.83755411743	11.3989140596	6.26268931023
2	1.13134757306	0.265075490479	-0.203016882839	1.14927515241	-0.105396398144	0.671155553593	0.056364875044	-0.180539718854	-0.0191105368218	0.192931027568	-0.0721137521457	0.231929258444	0.0314695350025	-0.0571114235463	0.0233014891698	0.232185098817	-0.00131987462774	0.0467261643746	0.205525145076	-0.00832547026467	0.193705456136	-10.8274479423	5.0746455097	18.5664731707
3	1.13155322075	-0.247971367479	-0.53014876336	1.00760556475	0.350425018194	0.996024542959	-0.031788299001	0.0384371113256	-0.0197112405971	0.0455008297271	-0.0028872213195	0.283655205418	0.0434448702855	-0.135705489503	0.0271807557936	0.168041368727	-0.00830453100322	0.00843322770185	0.179309451222	-0.0119673313559	0.198572057574	-14.3750518866	-4.08520910087	9.5922887343
4	4.62286148474	-1.54726982863	1.45668898357	2.03323898479	-0.777645782219	2.4830940479	0.0328806764278	0.269049650504	-0.185106953449	-0.302590563955	-0.108849651589	0.293106575777	0.277562576546	-0.0505366422549	0.129730922396	0.249990814787	-0.022170547186	-0.015219418729	0.191359901769	-0.0384080437686	0.210551034846	-11.2313553913	-14.4688159757	-0.150632637975
5	1.08223545033	0.42905618243	0.329937229807	0.972435843849	0.283651893871	0.668287131092	0.00831065508052	0.0651289043271	-0.080623194717	-0.115563868592	0.0274027075911	-0.0191690041594	0.0235187859537	0.0114505382214	-0.0257977397252	0.174435679947	0.00566776623924	0.00645967755838	0.149528583988	-0.0102052575083	0.131749319411	3.01226246784	-4.14681514209	1.75526579533
6	1.5761343871	0.18219982797	0.339884005268	2.04328551396	0.614175143012	2.37997425749	0.0391829983676	0.0750812316178	0.0547241715569	-0.0281627386098	0.0268614172416	-0.131128490774	0.00610268421039	0.224883748688	-0.0622536928085	0.137338566981	0.00132757076767	0.0175015824775	0.157864061561	-0.0169066182698	0.150957565145	15.1639839239	4.50399983906	7.95738208094
7	1.05570905159	0.227173838049	0.472492320454	1.53063267029	0.643081127612	2.29434348355	0.029077077287	-0.0146490646016	-0.0351660902629	0.0512710738918	-0.0463635493553	-0.0870275003757	0.0347717494904	-0.0356077124916	0.00222211965851	0.132708805043	-0.00405198057577	-0.00657083225012	0.122876718493	0.0068929522311	0.146086238996	13.8281896477	4.79060321788	36.935832588
8	1.1598389248	-0.114730531946	0.0161623930566	0.701977563952	0.0600345743494	0.342765944379	0.0290367776678	0.0596111714854	0.0642337551025	-0.0318305240915	-0.0518620619167	0.102819338584	-0.0321621592718	-0.0476068804347	0.0234793794404	0.133328154337	-0.000235038457144	-0.0143805644504	0.15940395964	-0.0091835337952	0.160161852818	-5.06677188055	11.6518646279	35.0456185624
9	1.36325388032	-0.0173655208816	0.611802246143	1.12954555151	-0.208337121751	0.743403173716	0.0157273777561	-0.11644865964	-0.00303437046019	0.080300011753	-0.0358632051589	0.202958717246	0.0336670715164	-0.0905839796235	0.0355357481019	0.115440624098	-0.0141406775143	0.0108218981636	0.157593219722	-0.0318912939806	0.140420613605	-8.07430422884	13.85070418	44.1288992345
10	1.31480395999	0.505807249223	0.0345155655887	1.15592484939	0.0169188816518	0.414840097251	0.0261140142877	-0.105620911312	0.171046741022	0.0445643341166	-0.0727673025554	0.143557930227	-0.0376025607136	-0.0382143564856	0.0407170055661	0.143944447554	0.00148349321174	0.000871758310368	0.163306069951	-0.0392825156945	0.163063739331	-0.809184160929	25.310815252	44.7616129953
11	1.69245844995	0.823676106125	-1.23107017433	1.11999447256	-0.649947543388	2.89647503466	0.00495195090347	-0.18320285813	0.180713341012	0.171330987825	-0.0889754664782	0.0725921789901	-0.00320858279433	0.000237951262289	0.054278028764	0.177895664403	0.00332464871899	0.00430563557763	0.199311848232	-0.048797703563	0.189402664424	5.33372462965	30.8585340236	49.9323384241
12	0.630985330415	-0.000529934483609	-0.0593358998307	0.915706462203	0.520475218399	0.980733762895	0.024826094814	-0.00782636622661	0.00393237391166	-0.00947777880921	-0.00176296648273	-0.0556257562676	-0.0436355401822	0.0266186391523	-0.0195798546074	0.115800335947	-0.00151671279289	-0.0084451804427	0.124502101063	-0.00473933816998	0.130629930272	17.247995514	17.2977177302	37.8686783874

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	chain 'A' and (resid 3 through 45 )
2	X-RAY DIFFRACTION	2	chain 'A' and (resid 46 through 100 )
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 101 through 181 )
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 182 through 196 )
5	X-RAY DIFFRACTION	5	chain 'A' and (resid 197 through 270 )
6	X-RAY DIFFRACTION	6	chain 'A' and (resid 271 through 303 )
7	X-RAY DIFFRACTION	7	chain 'B' and (resid 3 through 45 )
8	X-RAY DIFFRACTION	8	chain 'B' and (resid 46 through 100 )
9	X-RAY DIFFRACTION	9	chain 'B' and (resid 101 through 130 )
10	X-RAY DIFFRACTION	10	chain 'B' and (resid 131 through 181 )
11	X-RAY DIFFRACTION	11	chain 'B' and (resid 182 through 196 )
12	X-RAY DIFFRACTION	12	chain 'B' and (resid 197 through 303 )

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