PDB-5hht: Crystal structure of E. coli transketolase triple variant Ser385T...

ID or keywords:

Entry

Database: PDB / ID: 5hht

Title

Crystal structure of E. coli transketolase triple variant Ser385Tyr/Asp469Thr/Arg520Gln

Components

Transketolase 1

Keywords

TRANSFERASE /

thiamin diphosphate / engineered variant /

pentose phosphate pathway

Function / homology

Function and homology information

transketolase /

transketolase activity / pentose-phosphate shunt, non-oxidative branch / pentose-phosphate shunt /

thiamine pyrophosphate binding / manganese ion binding / magnesium ion binding / protein homodimerization activity /

cytosol
Similarity search - Function

Biological species

Escherichia coli K-12 (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 1.5 Å

Authors

Dai, S. / Tittmann, K.

Funding support

Germany, 1items

Citation

Journal: Sci Rep / Year: 2016
Title: Structural Analysis of an Evolved Transketolase Reveals Divergent Binding Modes.
Authors: Affaticati, P.E. / Dai, S.B. / Payongsri, P. / Hailes, H.C. / Tittmann, K. / Dalby, P.A.

History

Deposition	Jan 11, 2016	Deposition site: RCSB / Processing site: PDBE
Revision 1.0	Nov 2, 2016	Provider: repository / Type: Initial release
Revision 1.1	Sep 6, 2017	Group: Author supporting evidence / Data collection / Category: diffrn_source / pdbx_audit_support Item: _diffrn_source.pdbx_synchrotron_site / _pdbx_audit_support.funding_organization
Revision 2.0	Aug 4, 2021	Group: Atomic model / Data collection ...Atomic model / Data collection / Derived calculations / Non-polymer description / Structure summary Category: atom_site / chem_comp ...atom_site / chem_comp / entity / pdbx_entity_nonpoly / pdbx_nonpoly_scheme / pdbx_struct_conn_angle / struct_conn / struct_site / struct_site_gen Item: _atom_site.B_iso_or_equiv / _atom_site.Cartn_x ..._atom_site.B_iso_or_equiv / _atom_site.Cartn_x / _atom_site.Cartn_y / _atom_site.Cartn_z / _atom_site.auth_atom_id / _atom_site.auth_comp_id / _atom_site.label_atom_id / _atom_site.label_comp_id / _chem_comp.formula / _chem_comp.formula_weight / _chem_comp.id / _chem_comp.mon_nstd_flag / _chem_comp.name / _chem_comp.type / _entity.formula_weight / _entity.pdbx_description / _pdbx_entity_nonpoly.comp_id / _pdbx_entity_nonpoly.name / _pdbx_nonpoly_scheme.mon_id / _pdbx_nonpoly_scheme.pdb_mon_id / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _struct_conn.pdbx_dist_value / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_site.details / _struct_site.pdbx_auth_comp_id / _struct_site_gen.auth_comp_id / _struct_site_gen.label_comp_id
Revision 2.1	May 8, 2024	Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2 Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	5hht.cif.gz	538.5 KB	Display	PDBx/mmCIF format
PDB format	pdb5hht.ent.gz	432.1 KB	Display	PDB format
PDBx/mmJSON format	5hht.json.gz		Tree view	PDBx/mmJSON format
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Validation report

Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/hh/5hht ftp://data.pdbj.org/pub/pdb/validation_reports/hh/5hht	HTTPS FTP

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Related structure data

Similar structure data	3m49-d 4c7v-1 5i4i-1 5xsm-1 3upt-d 2r5n-d 1tka-d 5xtv-1 5xt0-1 6tj9-d 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	#153 - Sep 2012 Pyruvate Dehydrogenase Complex similarity (8) #263 - Nov 2021 Acetohydroxyacid Synthase similarity (2) #154 - Oct 2012 Citric Acid Cycle similarity (4)

Deposited unit

A: Transketolase 1

B: Transketolase 1

hetero molecules

149 kDa, 2 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)	89.944, 102.044, 133.076
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	19
Space group name H-M	P2₁2₁2₁

#1: Protein	Transketolase 1 / / TK 1 Mass: 73154.547 Da / Num. of mol.: 2 / Mutation: S385Y, D469T, R520Q Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K12 / Gene: tktA, tkt, b2935, JW5478 / Production host: Escherichia coli (E. coli) / References: UniProt: P27302, transketolase
#2: Chemical	... ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL / Ethylene glycol Mass: 62.068 Da / Num. of mol.: 36 / Source method: obtained synthetically / Formula: C₂H₆O₂
#3: Chemical	ChemComp-CA / CALCIUM ION Mass: 40.078 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Ca
#4: Chemical	ChemComp-TPP / THIAMINE DIPHOSPHATE / Thiamine pyrophosphate Mass: 425.314 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₁₂H₁₉N₄O₇P₂S
#5: Water	ChemComp-HOH / water / Water Mass: 18.015 Da / Num. of mol.: 1750 / Source method: isolated from a natural source / Formula: H₂O

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Experiment

Experiment	Method: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

Crystal	Density Matthews: 2.09 Å³/Da / Density % sol: 41.06 %
Crystal grow	Temperature: 281 K / Method: vapor diffusion, hanging drop / pH: 7.9 Details: The protein as apo-enzyme was concentrated to 16-20 mg/ml in 50 mM Gly-Gly buffer, pH 7.9. Afterwards 5 mM ThDP and 5 mM CaCl2 were added to the protein solution. Protein solution was mixed ...Details: The protein as apo-enzyme was concentrated to 16-20 mg/ml in 50 mM Gly-Gly buffer, pH 7.9. Afterwards 5 mM ThDP and 5 mM CaCl2 were added to the protein solution. Protein solution was mixed at 1+1 ratio with a reservoir solution containing 17-22 % (w/v) PEG 6000, 2 % glycerol, 50 mM Gly-Gly buffer, pH 7.9. PH range: 7.9

Crystal

Density Matthews: 2.09 Å³/Da / Density % sol: 41.06 %

Crystal grow

Temperature: 281 K / Method: vapor diffusion, hanging drop / pH: 7.9
Details: The protein as apo-enzyme was concentrated to 16-20 mg/ml in 50 mM Gly-Gly buffer, pH 7.9. Afterwards 5 mM ThDP and 5 mM CaCl2 were added to the protein solution. Protein solution was mixed ...
PH range: 7.9

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Data collection

Diffraction

Mean temperature: 100 K

Diffraction source

Source:

SYNCHROTRON / Site:

MAX II

/ Beamline: I911-3 / Wavelength: 1 Å

Detector

Type: MARMOSAIC 225 mm CCD / Detector: CCD / Date: Feb 26, 2015

Radiation

Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray

Radiation wavelength

Wavelength: 1 Å / Relative weight: 1

Reflection

Resolution: 1.5→44.38 Å / Num. obs: 190942 / % possible obs: 97.7 % / Observed criterion σ(I): -3 / Redundancy: 4.2 % / R_merge F obs: 0.999 / R_merge(I) obs: 0.045 / R_rim(I) all: 0.051 / Χ²: 0.978 / Net I/σ(I): 22.73 / Num. measured all: 797177

Reflection shell

Diffraction-ID: 1 / Rejects: 0

Resolution (Å)	Highest resolution (Å)	R_merge F obs	R_merge(I) obs	Mean I/σ(I) obs	Num. measured obs	Num. possible	Num. unique obs	R_rim(I) all	% possible all
1.5-1.6		0.942	0.201	7.45	138238	33876	33781	0.232	99.7
1.6-1.8		0.977	0.126	11.22	194873	47317	47118	0.145	99.6
1.8-2		0.993	0.07	18.23	125381	30319	29985	0.081	98.9
2-2.5		0.998	0.041	28.67	165351	39983	38975	0.047	97.5
2.5-2.9		0.999	0.03	37.38	62305	15175	14485	0.034	95.5
2.9-3		0.999	0.027	41.24	10705	2623	2482	0.031	94.6
3-3.68		0.999	0.023	47.51	45980	11274	10577	0.026	93.8
3.68-4.11		0.999	0.021	56.52	15354	3803	3513	0.023	92.4
4.11-4.54		0.999	0.019	59.36	10140	2505	2300	0.022	91.8
4.54-8		0.999	0.019	58.49	23860	5997	5410	0.022	90.2
8-14		0.999	0.019	61.86	4172	1142	967	0.021	84.7
14-17		0.999	0.02	61.58	408	129	100	0.023	77.5
17-33		0.999	0.02	52.21	388	143	105	0.023	73.4
33-50		0.998	0.03	12.12	22	18	9	0.042	50
	50					8

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Processing

Software

Name	Version	Classification
PHENIX	1.9_1692	refinement
XSCALE		data scaling
PDB_EXTRACT	3.15	data extraction
XDS		data reduction
PHENIX		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 1.5→44.379 Å / SU ML: 0.1 / Cross valid method: FREE R-VALUE / σ(F): 1.35 / Phase error: 13.08 / Stereochemistry target values: ML

	R_factor	Num. reflection	% reflection
R_free	0.1569	9625	5.04 %
R_work	0.1258	-	-
obs	0.1274	190942	97.69 %

Solvent computation

Shrinkage radii: 1.1 Å / VDW probe radii: 1.3 Å / Solvent model: FLAT BULK SOLVENT MODEL

Refinement step

Cycle: LAST / Resolution: 1.5→44.379 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	10204	0	198	1750	12152

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.013	10810
X-RAY DIFFRACTION	f_angle_d	1.573	14633
X-RAY DIFFRACTION	f_dihedral_angle_d	14.001	3965
X-RAY DIFFRACTION	f_chiral_restr	0.069	1559
X-RAY DIFFRACTION	f_plane_restr	0.008	1908

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 (%)
1.5-1.517	0.1899	344	0.1509	6036	X-RAY DIFFRACTION	100
1.517-1.5349	0.1777	329	0.1429	6115	X-RAY DIFFRACTION	100
1.5349-1.5536	0.1853	306	0.1412	6127	X-RAY DIFFRACTION	100
1.5536-1.5733	0.1815	305	0.1395	6162	X-RAY DIFFRACTION	100
1.5733-1.594	0.1824	323	0.1372	6104	X-RAY DIFFRACTION	100
1.594-1.6158	0.1689	319	0.1283	6122	X-RAY DIFFRACTION	100
1.6158-1.6389	0.1575	331	0.1313	6134	X-RAY DIFFRACTION	100
1.6389-1.6634	0.1746	342	0.1302	6092	X-RAY DIFFRACTION	100
1.6634-1.6894	0.1707	332	0.1294	6084	X-RAY DIFFRACTION	100
1.6894-1.7171	0.1657	341	0.1273	6092	X-RAY DIFFRACTION	100
1.7171-1.7467	0.1542	350	0.1252	6111	X-RAY DIFFRACTION	100
1.7467-1.7784	0.1453	315	0.1195	6111	X-RAY DIFFRACTION	99
1.7784-1.8126	0.1508	303	0.1223	6151	X-RAY DIFFRACTION	99
1.8126-1.8496	0.1477	341	0.1224	6094	X-RAY DIFFRACTION	99
1.8496-1.8899	0.1564	328	0.1244	6103	X-RAY DIFFRACTION	99
1.8899-1.9338	0.1635	324	0.1218	6057	X-RAY DIFFRACTION	99
1.9338-1.9822	0.1493	317	0.1234	6102	X-RAY DIFFRACTION	99
1.9822-2.0358	0.143	335	0.1205	6036	X-RAY DIFFRACTION	98
2.0358-2.0957	0.1579	335	0.1224	6070	X-RAY DIFFRACTION	98
2.0957-2.1633	0.1501	334	0.119	6027	X-RAY DIFFRACTION	98
2.1633-2.2406	0.1531	333	0.1187	6028	X-RAY DIFFRACTION	98
2.2406-2.3303	0.1644	315	0.1214	5998	X-RAY DIFFRACTION	97
2.3303-2.4364	0.1439	298	0.1248	6040	X-RAY DIFFRACTION	97
2.4364-2.5648	0.1465	329	0.1253	5954	X-RAY DIFFRACTION	96
2.5648-2.7255	0.1641	291	0.1267	5966	X-RAY DIFFRACTION	96
2.7255-2.9359	0.1588	327	0.1302	5913	X-RAY DIFFRACTION	95
2.9359-3.2313	0.1549	303	0.1276	5908	X-RAY DIFFRACTION	94
3.2313-3.6987	0.1528	301	0.1211	5872	X-RAY DIFFRACTION	93
3.6987-4.6591	0.142	293	0.1131	5880	X-RAY DIFFRACTION	92
4.6591-44.3983	0.1589	281	0.1405	5828	X-RAY DIFFRACTION	88

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	0.035	0.0084	0.0072	0.0175	-0.0038	0.0183	-0.0357	-0.0286	-0.0285	0.0098	0.0096	0.0019	0.0305	0.0123	-0	0.0476	0.0064	0.0114	0.0434	0.0075	0.0555	24.9794	-27.2737	16.3759
2	0.0164	-0.0272	-0.0073	0.0488	0.0252	0.0083	-0.0181	0.0151	-0.025	-0.0095	0.007	-0.019	0.0096	0.0071	-0.0003	0.0249	0.0068	0.0015	0.0408	0.0018	0.0483	30.9079	-13.0225	8.7089
3	0.0042	0.0027	-0.0028	0.0106	0.0072	0.0101	-0.0052	-0.0055	-0.0125	-0.0425	-0.012	-0.0208	0.0237	0.0058	-0	0.0499	0.0088	0.0063	0.041	-0.0026	0.0456	28.2322	-19.5551	-4.7085
4	0.0547	-0.0182	-0.0308	0.0204	-0.0003	0.0354	-0.0425	0.0301	-0.0513	-0.0308	0.0207	-0.007	0.0418	-0.0095	-0.0079	0.044	0.005	0.0125	0.0383	0.0011	0.0744	19.9446	-32.5515	8.4215
5	0.0517	0.0302	-0.0307	0.1043	0.0666	0.0502	0.0041	-0.0139	-0.0075	0.0247	0.0154	-0.0377	-0.009	0.0152	0.011	0.0241	0.0025	-0.0114	0.0364	-0.0022	0.0404	36.4505	2.9461	18.0824
6	0.0713	-0.0114	0.043	0.0595	0.0031	0.0311	-0.0019	-0.0238	-0.0029	0.0417	0.0085	0.0019	-0.0097	0.0097	-0	0.0436	-0.0025	0.0041	0.0333	-0.0054	0.0296	21.6849	17.4006	33.0785
7	0.0065	0.006	0.0006	0.0045	0.0062	0.0044	-0.016	0.0179	0.0498	-0.039	0.0067	0.0308	-0.0449	-0.0247	-0	0.0806	0.0024	0.0003	0.0447	-0.0067	0.0446	11.772	28.0935	21.3129
8	0.1198	0.0568	-0.0869	0.1055	-0.0382	0.1302	-0.0013	0.0233	0.0186	-0.008	0.011	0.0154	-0.011	-0.0134	0.0007	0.0237	0.0013	-0.0059	0.0276	0.0061	0.0228	6.7576	7.8591	0.6407
9	0.028	0.012	-0.0032	0.0062	0.0031	0.0096	-0.044	-0.0597	-0.0341	0.0482	0.0237	0.001	0.0049	0.0366	-0	0.0701	0.0112	0.0137	0.0617	-0.0001	0.0494	4.5439	-2.5594	36.4306

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	chain 'A' and (resid 2 through 99 )
2	X-RAY DIFFRACTION	2	chain 'A' and (resid 100 through 192 )
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 193 through 233 )
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 234 through 296 )
5	X-RAY DIFFRACTION	5	chain 'A' and (resid 297 through 477 )
6	X-RAY DIFFRACTION	6	chain 'A' and (resid 478 through 630 )
7	X-RAY DIFFRACTION	7	chain 'A' and (resid 631 through 663 )
8	X-RAY DIFFRACTION	8	chain 'B' and (resid 2 through 615 )
9	X-RAY DIFFRACTION	9	chain 'B' and (resid 616 through 668 )

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