PDB-2ay0: Structure of the Lys9Met mutant of the E. coli Proline Utilizatio...

ID or keywords:

Entry

Database: PDB / ID: 2ay0

Title

Structure of the Lys9Met mutant of the E. coli Proline Utilization A (PutA) DNA-binding domain.

Components

Bifunctional putA protein

Keywords

DNA BINDING PROTEIN / PutA / ribbon-helix-helix / dna-binding domain / proline catabolism / proline utilization A

Function / homology

Function and homology information

proline dehydrogenase / proline dehydrogenase activity / L-glutamate gamma-semialdehyde dehydrogenase / 1-pyrroline-5-carboxylate dehydrogenase activity / proline catabolic process to glutamate / proline biosynthetic process / DNA-binding transcription repressor activity / cis-regulatory region sequence-specific DNA binding / cytoplasmic side of plasma membrane / flavin adenine dinucleotide binding ...
Similarity search - Function

Biological species

Escherichia coli (E. coli)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MAD / Resolution: 2.1 Å

Authors

Larson, J.D. / Schuermann, J.P. / Zhou, Y. / Jenkins, J.L. / Becker, D.F. / Tanner, J.J.

Citation

Journal: Protein Sci. / Year: 2006
Title: Crystal structures of the DNA-binding domain of Escherichia coli proline utilization A flavoprotein and analysis of the role of Lys9 in DNA recognition.
Authors: Larson, J.D. / Jenkins, J.L. / Schuermann, J.P. / Zhou, Y. / Becker, D.F. / Tanner, J.J.

History

Deposition	Sep 6, 2005	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Aug 15, 2006	Provider: repository / Type: Initial release
Revision 1.1	May 1, 2008	Group: Version format compliance
Revision 1.2	Jul 13, 2011	Group: Advisory / Version format compliance
Revision 1.3	Oct 20, 2021	Group: Database references / Derived calculations / Category: database_2 / struct_ref_seq_dif / struct_site Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_ref_seq_dif.details / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id
Revision 1.4	Feb 14, 2024	Group: Data collection / Category: chem_comp_atom / chem_comp_bond

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	2ay0.cif.gz	65 KB	Display	PDBx/mmCIF format
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Validation report

Summary document	2ay0_validation.pdf.gz	476.2 KB	Display	wwPDB validaton report
Full document	2ay0_full_validation.pdf.gz	479.8 KB	Display
Data in XML	2ay0_validation.xml.gz	11.7 KB	Display
Data in CIF	2ay0_validation.cif.gz	16.2 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/ay/2ay0 ftp://data.pdbj.org/pub/pdb/validation_reports/ay/2ay0	HTTPS FTP

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Related structure data

Related structure data	2gpeC C: citing same article (ref.)
Similar structure data	4o6x-1 5d13-4 6v84-1 1fkg-d 2wa9-1 4nmv-2 5cax-3 2o71-d 5aeo-1 6h8o-2 Search similar-shape structures by Omokage search (details) 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

Related structure data

2gpeC

C: citing same article (ref.)

Similar structure data

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Links

PDB pages	PDBj / wwPDB / NCBI
Related items in Molecule of the Month	#127 - Jul 2010 Crystallins similarity (7) #89 - May 2007 Aconitase and Iron Regulatory Protein 1 similarity (7)

Deposited unit

A: Bifunctional putA protein

B: Bifunctional putA protein

C: Bifunctional putA protein

D: Bifunctional putA protein

E: Bifunctional putA protein

F: Bifunctional putA protein

hetero molecules

40.9 kDa, 6 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: Bifunctional putA protein

B: Bifunctional putA protein

hetero molecules

defined by author&software
13.6 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

C: Bifunctional putA protein

D: Bifunctional putA protein

hetero molecules

defined by author&software
13.6 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

3

E: Bifunctional putA protein

F: Bifunctional putA protein

hetero molecules

defined by author&software
13.6 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

Unit cell

Length a, b, c (Å)	72.070, 91.494, 69.606
Angle α, β, γ (deg.)	90.00, 119.21, 90.00
Int Tables number	5
Space group name H-M	C121

Components on special symmetry positions

ID	Model	Components
1	1	B-71- HOH

Details

The biological assemble is a dimer. There are 3 biological dimers in the asymmetric unit: A/B, C/D and E/F.

#1: Protein	Bifunctional putA protein Mass: 6788.644 Da / Num. of mol.: 6 / Fragment: residues 1-52 / Mutation: K9M Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: putA, poaA / Plasmid: pET23b / Production host: Escherichia coli (E. coli) / Strain (production host): BL21DE3 pLysS Keywords: DNA-binding domain of E. coli Proline utilization A References: UniProt: P09546
#2: Chemical	ChemComp-CL / CHLORIDE ION Mass: 35.453 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Cl
#3: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 67 / 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: 2.46 Å³/Da / Density % sol: 49.96 %
Crystal grow	Temperature: 295 K / Method: vapor diffusion, hanging drop Details: 1.7-3.0 M NaCl, 5 mM dithiothreitol, pH 3.0-5.5, VAPOR DIFFUSION, HANGING DROP, temperature 295K

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Data collection

Diffraction

ID	Mean temperature (K)	Crystal-ID
1	173	1
2		1
3		1

Diffraction source

Source:

SYNCHROTRON / Site:

ALS

/ Beamline: 4.2.2 / Wavelength: 0.987121, 0.979553, 0.979144

Detector

Type: CUSTOM-MADE / Detector: CCD / Date: Dec 8, 2004

Radiation

Monochromator: ALS beamline 4.2.2 / Protocol: MAD / Monochromatic (M) / Laue (L): M / Scattering type: x-ray

Radiation wavelength

ID	Wavelength (Å)	Relative weight
1	0.987121	1
2	0.979553	1
3	0.979144	1

Reflection

Av σ(I) over netI: 12.5 / Number: 25 / R_merge(I) obs: 0.079 / Χ²: 0.99 / D res high: 2 Å / D res low: 36.53 Å / Num. obs: 26576 / % possible obs: 100 / Redundancy: 7.47 % / Rejects: 1501

ID
1
2
3

Diffraction reflection shell

Highest resolution (Å)	Lowest resolution (Å)	% possible obs (%)	ID	R_merge(I) obs	Chi squared	Rejects
4.31	36.53	99.6	1	0.06	1.13	652
3.42	4.31	100	1	0.053	0.9	122
2.99	3.42	100	1	0.066	0.89	102
2.71	2.99	100	1	0.096	0.93	105
2.52	2.71	100	1	0.15	0.96	126
2.37	2.52	100	1	0.203	0.99	100
2.25	2.37	100	1	0.267	1	65
2.15	2.25	100	1	0.357	1	80
2.07	2.15	100	1	0.426	1.03	78
2	2.07	100	1	0.52	1.04	71
4.31	36.53	99.6	2	0.06	1.13	652
3.42	4.31	100	2	0.053	0.9	122
2.99	3.42	100	2	0.066	0.89	102
2.71	2.99	100	2	0.096	0.93	105
2.52	2.71	100	2	0.15	0.96	126
2.37	2.52	100	2	0.203	0.99	100
2.25	2.37	100	2	0.267	1	65
2.15	2.25	100	2	0.357	1	80
2.07	2.15	100	2	0.426	1.03	78
2	2.07	100	2	0.52	1.04	71
4.31	36.53	99.6	3	0.06	1.13	652
3.42	4.31	100	3	0.053	0.9	122
2.99	3.42	100	3	0.066	0.89	102
2.71	2.99	100	3	0.096	0.93	105
2.52	2.71	100	3	0.15	0.96	126
2.37	2.52	100	3	0.203	0.99	100
2.25	2.37	100	3	0.267	1	65
2.15	2.25	100	3	0.357	1	80
2.07	2.15	100	3	0.426	1.03	78
2	2.07	100	3	0.52	1.04	71

Reflection

Resolution: 2→60.75 Å / Num. all: 26606 / Num. obs: 26606 / % possible obs: 100 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 7.47 % / R_merge(I) obs: 0.079 / Net I/σ(I): 12.5

Reflection shell

Resolution (Å)	R_merge(I) obs	Mean I/σ(I) obs	Diffraction-ID
2-2.07	0.52	3.7	1
2.07-2.15	0.426	4.3	1
2.15-2.25	0.357	4.9	1
2.25-2.37	0.267	6.2	1
2.37-2.52	0.203	7.7	1
2.52-2.71	0.15	10	1
2.71-2.99	0.096	14.3	1
2.99-3.42	0.066	20.5	1
3.42-4.31	0.053	27	1
4.31-36.53	0.06	26.4	1

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Phasing

Phasing

Method:

MAD

Phasing MAD set site

ID	Atom type symbol	B iso	Fract x	Fract y	Fract z	Occupancy
1	Se	60	0.34	0.5	0.155	0.928
2	Se	56.677	0.366	0.308	0.411	0.956
3	Se	51.945	0.059	0.5	0.063	0.794
4	Se	60	0.83	0.1	0.267	1.028
5	Se	48.575	0.386	0.156	0.073	0.746
6	Se	60	0.351	0.202	0.294	0.984
7	Se	60	0.501	0.433	0.284	0.929
8	Se	60	0.557	0.124	0.086	1.012
9	Se	57.582	0.414	0.489	0.374	0.845
10	Se	60	0.471	0.037	0.162	0.796
11	Se	60	0.51	0.136	0.4	0.967
12	Se	60	0.531	0.265	0.427	0.777

Phasing dm

FOM : 0.54 / FOM acentric: 0.54 / FOM centric: 0.59 / Reflection: 26555 / Reflection acentric: 25713 / Reflection centric: 842

Phasing dm shell

Resolution (Å)	FOM	FOM acentric	FOM centric	Reflection	Reflection acentric	Reflection centric
5.7-19.847	0.97	0.97	0.92	1148	1052	96
3.6-5.7	0.95	0.96	0.93	3555	3392	163
2.9-3.6	0.86	0.86	0.77	4467	4313	154
2.5-2.9	0.64	0.65	0.58	4486	4357	129
2.1-2.5	0.34	0.35	0.28	7981	7785	196
2-2.1	0.09	0.09	0.08	4918	4814	104

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Processing

Software

Name	Version	Classification
d*TREK	9.2LDz	data scaling
SOLVE	2.06	phasing
RESOLVE	2.06	phasing
REFMAC		refinement
PDB_EXTRACT	1.7	data extraction
Blu-Ice		data collection

Refinement

Method to determine structure:

MAD / Resolution: 2.1→60.75 Å / Cor.coef. F_o:F_c: 0.951 / Cor.coef. F_o:F_c free: 0.93 / SU B: 8.155 / SU ML: 0.123 / TLS residual ADP flag: LIKELY RESIDUAL / Cross valid method: THROUGHOUT / ESU R: 0.175 / ESU R Free: 0.162 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS

	R_factor	Num. reflection	% reflection	Selection details
R_free	0.2418	1153	5 %	RANDOM
R_work	0.204	-	-	-
obs	0.20597	21873	99.88 %	-
all	-	26606	-	-

Solvent computation

Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK

Displacement parameters

B_iso mean: 38.245 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	0.61 Å²	0 Å²	-0.67 Å²
2-	-	0.11 Å²	0 Å²
3-	-	-	-1.38 Å²

Refinement step

Cycle: LAST / Resolution: 2.1→60.75 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	2072	0	6	67	2145

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.011	0.022	2102
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.02	1997
X-RAY DIFFRACTION	r_angle_refined_deg	1.139	1.951	2842
X-RAY DIFFRACTION	r_angle_other_deg	0.774	3	4596
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	4.667	5	255
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	32.346	22.727	88
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	15.652	15	387
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	14.232	15	20
X-RAY DIFFRACTION	r_chiral_restr	0.069	0.2	343
X-RAY DIFFRACTION	r_gen_planes_refined	0.004	0.02	2253
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	425
X-RAY DIFFRACTION	r_nbd_refined	0.219	0.2	477
X-RAY DIFFRACTION	r_nbd_other	0.167	0.2	1913
X-RAY DIFFRACTION	r_nbtor_refined	0.171	0.2	1016
X-RAY DIFFRACTION	r_nbtor_other	0.082	0.2	1276
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.147	0.2	58
X-RAY DIFFRACTION	r_xyhbond_nbd_other
X-RAY DIFFRACTION	r_metal_ion_refined
X-RAY DIFFRACTION	r_metal_ion_other
X-RAY DIFFRACTION	r_symmetry_vdw_refined	0.253	0.2	15
X-RAY DIFFRACTION	r_symmetry_vdw_other	0.167	0.2	48
X-RAY DIFFRACTION	r_symmetry_hbond_refined	0.097	0.2	6
X-RAY DIFFRACTION	r_symmetry_hbond_other
X-RAY DIFFRACTION	r_symmetry_metal_ion_refined
X-RAY DIFFRACTION	r_symmetry_metal_ion_other
X-RAY DIFFRACTION	r_mcbond_it	0.732	1.5	1414
X-RAY DIFFRACTION	r_mcbond_other	0.137	1.5	536
X-RAY DIFFRACTION	r_mcangle_it	1.007	2	2104
X-RAY DIFFRACTION	r_scbond_it	1.637	3	877
X-RAY DIFFRACTION	r_scangle_it	2.473	4.5	738
X-RAY DIFFRACTION	r_rigid_bond_restr
X-RAY DIFFRACTION	r_sphericity_free
X-RAY DIFFRACTION	r_sphericity_bonded

LS refinement shell

Resolution: 2.1→2.155 Å / Total num. of bins used: 20

	R_factor	Num. reflection	% reflection
R_free	0.295	72	-
R_work	0.234	1632	-
obs	-	-	100 %

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	4.2033	-1.5858	3.467	4.7459	-2.3981	10.6318	0.211	-0.0152	-0.1848	-0.2905	-0.1569	-0.0056	0.4869	0.2704	-0.0541	-0.0218	0.086	-0.0081	-0.1271	0.0013	-0.1213	12.313	42.738	10.814
2	3.2266	-0.0055	0.3153	8.3023	-5.6434	10.4517	0.0677	0.0755	0.1329	-0.3419	-0.1229	0.2529	-0.1346	-0.0792	0.0552	-0.1894	0.0813	-0.035	-0.0783	-0.0307	-0.1399	13.466	39.894	20.035
3	4.0697	-0.2911	-0.0579	8.4073	2.0508	6.4585	0.0421	0.1763	-0.1729	-0.1328	-0.0365	-0.0977	0.2191	0.3264	-0.0056	-0.1414	0.1121	0.0344	-0.0346	-0.0133	-0.1549	24.388	27.248	19.506
4	6.7255	-1.0593	-2.0368	7.0391	2.5503	4.995	-0.0335	0.2613	0.0415	-0.0258	-0.1426	0.2568	0.0541	-0.2278	0.1761	-0.0313	0.0788	-0.0046	-0.0635	-0.0309	-0.1241	21.218	21.201	14.192
5	3.8279	-2.07	-0.3556	4.5871	1.9827	5.4508	-0.1668	-0.3015	-0.05	0.4077	0.0289	0.1608	0.3796	0.0608	0.138	0.0041	0.064	0.0178	-0.0596	-0.0189	-0.1082	30.826	7.758	14.961
6	7.2346	0.283	1.1067	3.2109	0.2305	5.1189	-0.1948	-0.298	0.3784	0.1298	0.0096	-0.0195	-0.225	0.1103	0.1852	0.003	0.079	-0.0165	-0.142	-0.0493	-0.1298	38.927	10.679	11.72

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Auth asym-ID	Label asym-ID	Auth seq-ID	Label seq-ID
1	X-RAY DIFFRACTION	1	A	A	3 - 45	3 - 45
2	X-RAY DIFFRACTION	2	B	B	3 - 44	3 - 44
3	X-RAY DIFFRACTION	3	C	C	2 - 45	2 - 45
4	X-RAY DIFFRACTION	4	D	D	3 - 47	3 - 47
5	X-RAY DIFFRACTION	5	E	E	2 - 45	2 - 45
6	X-RAY DIFFRACTION	6	F	F	3 - 45	3 - 45

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