PDB-4f5k: Substrate Specificity Conversion of Aspartate Aminotransferase to...

ID or keywords:

Entry

Database: PDB / ID: 4f5k

Title

Substrate Specificity Conversion of Aspartate Aminotransferase to Tyrosine Aminotransferase By The JANUS Algorithm: Chimera P6.

Components

Aspartate aminotransferase

Keywords

TRANSFERASE / Aminotransferase

Function / homology

Function and homology information

L-phenylalanine biosynthetic process from chorismate via phenylpyruvate / L-tyrosine-2-oxoglutarate transaminase activity / L-phenylalanine biosynthetic process / aspartate transaminase / L-aspartate:2-oxoglutarate aminotransferase activity / pyridoxal phosphate binding / protein homodimerization activity / identical protein binding / cytosol / cytoplasm
Similarity search - Function

Biological species

Escherichia coli (E. coli)

Method

X-RAY DIFFRACTION /

MOLECULAR REPLACEMENT / Resolution: 2.2 Å

Authors

Addington, T.A. / Fisher, A.J. / Toney, M.D.

Citation

Journal: J.Mol.Biol. / Year: 2013
Title: Janus: prediction and ranking of mutations required for functional interconversion of enzymes.
Authors: Addington, T.A. / Mertz, R.W. / Siegel, J.B. / Thompson, J.M. / Fisher, A.J. / Filkov, V. / Fleischman, N.M. / Suen, A.A. / Zhang, C. / Toney, M.D.

History

Deposition	May 13, 2012	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Feb 13, 2013	Provider: repository / Type: Initial release
Revision 1.1	Feb 27, 2013	Group: Database references
Revision 1.2	Apr 24, 2013	Group: Database references

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	4f5k.cif.gz	324.5 KB	Display	PDBx/mmCIF format
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Validation report

Summary document	4f5k_validation.pdf.gz	439.4 KB	Display	wwPDB validaton report
Full document	4f5k_full_validation.pdf.gz	449.2 KB	Display
Data in XML	4f5k_validation.xml.gz	38.5 KB	Display
Data in CIF	4f5k_validation.cif.gz	58.1 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/f5/4f5k ftp://data.pdbj.org/pub/pdb/validation_reports/f5/4f5k	HTTPS FTP

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Related structure data

Related structure data	4f5fC 4f5gC 4f5hC 4f5iC 4f5jC 4f5mC 4f5l C: citing same article (ref.)
Similar structure data	1g7w-1 4f5i-d 1cq6-1 1asf-1 1bqa-d 1asb-1 4f5j-d 4f5h-d 1aic-d 1g4x-1 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

Deposited unit

A: Aspartate aminotransferase

B: Aspartate aminotransferase

89.6 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)	59.300, 102.346, 138.514
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	19
Space group name H-M	P2₁2₁2₁

#1: Protein	Aspartate aminotransferase / AspAT / Transaminase A Mass: 44810.426 Da / Num. of mol.: 2 Mutation: I39V, N40D, I43V, L56M, N74T, I78L, I81L, T114S, S139T, S145A, V146I, I197A, F220I, F222I, A228G, Y254C, G259S, S283G, V385I Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Strain: K12 / Gene: AAT, aspC, b0928, JW0911 / Plasmid: pET45b / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: P00509, aspartate transaminase
#2: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 788 / Source method: isolated from a natural source / Formula: H₂O

#1: Protein

Aspartate aminotransferase / AspAT / Transaminase A

Mass: 44810.426 Da / Num. of mol.: 2
Mutation: I39V, N40D, I43V, L56M, N74T, I78L, I81L, T114S, S139T, S145A, V146I, I197A, F220I, F222I, A228G, Y254C, G259S, S283G, V385I
Source method: isolated from a genetically manipulated source
Source: (gene. exp.)

Escherichia coli (E. coli) / Strain: K12 / Gene: AAT, aspC, b0928, JW0911 / Plasmid: pET45b / Production host:

Escherichia coli (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: P00509, aspartate transaminase

#2: Water

ChemComp-HOH / water

Mass: 18.015 Da / Num. of mol.: 788 / 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: 2.34 Å³/Da / Density % sol: 47.55 %
Crystal grow	Temperature: 277 K / Method: vapor diffusion, hanging drop / pH: 4.8 Details: 8% PEG 4000, 0.1 M Sodium Acetate, pH 4.8, VAPOR DIFFUSION, HANGING DROP, temperature 277K

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Data collection

Diffraction	Mean temperature: 85 K
Diffraction source	Source: ROTATING ANODE / Type: RIGAKU RU300 / Wavelength: 1.54 Å
Detector	Type: BRUKER SMART 6000 / Detector: CCD / Date: Jul 27, 2010 / Details: Mirrors
Radiation	Monochromator: Osmic Mirrors / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 1.54 Å / Relative weight: 1
Reflection	Resolution: 2.2→100 Å / Num. all: 43677 / Num. obs: 43617 / % possible obs: 99.9 % / Observed criterion σ(F): -3 / Observed criterion σ(I): -3 / Redundancy: 5.2 % / B_iso Wilson estimate: 22.9 Å² / R_merge(I) obs: 0.0984 / R_sym value: 0.0984 / Net I/σ(I): 11.92
Reflection shell	Resolution: 2.2→2.3 Å / Redundancy: 4.24 % / R_merge(I) obs: 0.335 / Mean I/σ(I) obs: 3 / Num. unique all: 5359 / R_sym value: 0.335 / % possible all: 100

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Processing

Software

Name	Version	Classification
PROTEUM PLUS	PLUS	data collection
PHASER		phasing
PHENIX	(phenix.refine: 1.7.1_743)	refinement
PROTEUM PLUS	PLUS	data reduction
PROTEUM PLUS	PLUS	data scaling

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 2.2→42.087 Å / σ(F): 1.34

	R_factor	Num. reflection	% reflection
R_free	0.2268	2117	-
R_work	0.1809	-	-
obs	0.1831	43538	99.86 %

Refine analyze

Luzzati sigma a obs: 0.53 Å

Refinement step

Cycle: LAST / Resolution: 2.2→42.087 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	6172	0	0	788	6960

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.007	6321
X-RAY DIFFRACTION	f_angle_d	1.085	8579
X-RAY DIFFRACTION	f_dihedral_angle_d	13.415	2303
X-RAY DIFFRACTION	f_chiral_restr	0.07	950
X-RAY DIFFRACTION	f_plane_restr	0.004	1132

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 (%)
2.2-2.2512	0.3247	147	0.2197	2705	X-RAY DIFFRACTION	100
2.2512-2.3075	0.301	157	0.2199	2712	X-RAY DIFFRACTION	100
2.3075-2.3699	0.2646	147	0.2174	2721	X-RAY DIFFRACTION	100
2.3699-2.4396	0.2583	141	0.1955	2706	X-RAY DIFFRACTION	100
2.4396-2.5183	0.2352	140	0.1999	2739	X-RAY DIFFRACTION	100
2.5183-2.6083	0.2716	139	0.1928	2738	X-RAY DIFFRACTION	100
2.6083-2.7127	0.2586	147	0.1936	2727	X-RAY DIFFRACTION	100
2.7127-2.8362	0.2414	147	0.1865	2739	X-RAY DIFFRACTION	100
2.8362-2.9857	0.2381	117	0.1836	2783	X-RAY DIFFRACTION	100
2.9857-3.1727	0.2313	158	0.1825	2714	X-RAY DIFFRACTION	100
3.1727-3.4175	0.2072	130	0.1836	2779	X-RAY DIFFRACTION	100
3.4175-3.7613	0.2239	140	0.1637	2786	X-RAY DIFFRACTION	100
3.7613-4.305	0.1481	138	0.152	2786	X-RAY DIFFRACTION	100
4.305-5.4221	0.1932	143	0.1532	2823	X-RAY DIFFRACTION	99
5.4221-42.0945	0.22	126	0.1854	2963	X-RAY DIFFRACTION	99

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	0.1372	0.0518	0.0249	0.1956	0.1902	0.5086	0.0974	-0.1056	0.112	-0.0615	0.0464	-0.1005	0.1627	-0.1628	0.1018	0.1635	0.031	0.039	0.1622	-0.0008	0.1539	15.0604	31.4049	3.6686
2	0.0451	0.0254	0.2063	0.1293	-0.0254	0.3274	-0.009	-0.054	-0.043	0.0028	0.019	-0.0114	-0.0088	0.0954	-0.0101	0.1077	-0.0161	-0.0064	0.141	0.0021	0.1531	37.0449	49.8635	15.3773
3	0.2999	0.1555	0.0866	0.1092	0.0525	0.3039	-0.0472	0.0554	0.0238	-0.0553	-0.014	0.0816	-0.223	-0.0127	-0.0382	0.1834	0.0189	-0.0269	0.1146	0.0049	0.1573	20.9489	60.842	-0.2169
4	0.2623	0.0954	0.0184	0.2095	0.0515	0.6202	-0.0106	0.0057	-0.0226	-0.0417	-0.0084	0.0134	-0.0938	0.0733	-0.0078	0.1004	-0.0012	-0.0021	0.1185	-0.0018	0.1236	32.0192	48.5411	0.005
5	0.4445	0.1843	-0.1704	0.1687	-0.025	0.0969	0.0041	-0.1144	-0.0481	-0.1182	-0.0894	-0.0096	-0.2793	0.2937	-0.0257	0.2907	-0.0218	-0.0221	0.2082	0.0152	0.1553	32.4793	59.8922	35.306
6	0.0999	0.0548	-0.1253	0.1597	-0.0619	0.1126	0.0075	-0.0728	-0.0202	0.0011	-0.0286	-0.0654	-0.0213	0.1081	-0.0023	0.1121	0.0221	-0.0027	0.1922	0.0013	0.1605	37.7199	33.8797	24.5978
7	0.0024	0.0021	0.013	0.0392	-0.037	0.0723	-0.0099	0.0135	-0.095	-0.0012	-0.0628	-0.154	0.1526	-0.0252	-0.0016	0.2423	-0.0362	-0.0175	0.1281	-0.0051	0.223	18.6004	23.1247	24.4476
8	0.078	-0.0906	-0.0839	0.167	0.1981	0.4627	-0.051	-0.0181	0.0246	0.0403	-0.0005	0.0029	0.0857	-0.0688	-0.0579	0.13	-0.0157	0.0244	0.1109	-0.0003	0.1228	12.4475	38.3955	36.8113
9	0.178	-0.1961	-0.1353	0.1189	0.135	0.1613	0.0015	0.0827	0.0295	-0.0334	-0.0368	0.0109	0.1234	0.0489	-0.0051	0.1147	-0.0033	0.0003	0.1187	0.0119	0.1422	20.7996	34.4379	22.8372
10	0.0386	-0.0485	0.0282	0.0944	-0.0645	0.0404	-0.0464	-0.0925	-0.0342	0.1147	0.0903	-0.0996	0.1703	0.0454	-0.0186	0.1842	0.0379	-0.0285	0.1759	0.0327	0.1608	34.8179	30.7053	46.1669
11	0.105	0.1362	-0.0001	0.1452	-0.0077	0.1445	-0.059	-0.0548	0.1114	0.1405	0.032	0.0082	0.0155	0.1596	-0	0.2283	0.02	-0.0341	0.2164	-0.0331	0.1908	31.4422	46.8472	53.857

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	CHAIN A AND (RESSEQ 8:42)
2	X-RAY DIFFRACTION	2	CHAIN A AND (RESSEQ 43:127)
3	X-RAY DIFFRACTION	3	CHAIN A AND (RESSEQ 128:214)
4	X-RAY DIFFRACTION	4	CHAIN A AND (RESSEQ 215:406)
5	X-RAY DIFFRACTION	5	CHAIN B AND (RESSEQ 8:42)
6	X-RAY DIFFRACTION	6	CHAIN B AND (RESSEQ 43:81)
7	X-RAY DIFFRACTION	7	CHAIN B AND (RESSEQ 82:104)
8	X-RAY DIFFRACTION	8	CHAIN B AND (RESSEQ 105:244)
9	X-RAY DIFFRACTION	9	CHAIN B AND (RESSEQ 245:310)
10	X-RAY DIFFRACTION	10	CHAIN B AND (RESSEQ 311:342)
11	X-RAY DIFFRACTION	11	CHAIN B AND (RESSEQ 343:406)

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