PDB-4ezu: Crystal structure of the substrate binding domain of E.coli DnaK ...

Basic information

• Entry: Database: PDB / ID: 4ezu
• Title: Crystal structure of the substrate binding domain of E.coli DnaK in complex with PR-bombesin in space group I222

Components

• Chaperone protein DnaK
• Proline rich bombesin-related protein

• Keywords: CHAPERONE/PEPTIDE BINDING PROTEIN / chaperone / peptide binding / CHAPERONE-PEPTIDE BINDING PROTEIN complex

Function / homology

Function:

stress response to copper ion / sigma factor antagonist activity / : / protein unfolding / cellular response to unfolded protein / neuropeptide signaling pathway / heat shock protein binding / inclusion body / protein folding chaperone / ATP-dependent protein folding chaperone / ADP binding / unfolded protein binding / protein-folding chaperone binding / response to heat / protein refolding / protein-containing complex assembly / DNA replication / protein-containing complex / ATP hydrolysis activity / extracellular region / zinc ion binding / ATP binding / membrane / plasma membrane / cytoplasm / cytosol

Domain/homology:

Bombesin/neuromedin-B/ranatensin peptide family / Bombesin-like peptide / Bombesin-like peptides family signature. / Substrate Binding Domain Of Dnak; Chain:A; Domain 2 - #10 / Chaperone DnaK / Substrate Binding Domain Of DNAk; Chain A, domain 1 / Substrate Binding Domain Of DNAk; Chain A, domain 1 / Heat shock hsp70 proteins family signature 2. / Heat shock hsp70 proteins family signature 1. / Heat shock hsp70 proteins family signature 3. / Heat shock protein 70, conserved site / Heat shock protein 70kD, peptide-binding domain superfamily / Heat shock protein 70kD, C-terminal domain superfamily / Substrate Binding Domain Of Dnak; Chain:A; Domain 2 / Heat shock protein 70 family / Hsp70 protein / ATPase, nucleotide binding domain / Up-down Bundle / Sandwich / Mainly Beta / Mainly Alpha

Component:

Chaperone protein DnaK / Proline rich bombesin-related protein

• Biological species: Escherichia coli (E. coli); Bombina maxima (large-webbed bell toad)
• Method: X-RAY DIFFRACTION / SYNCHROTRON / PDB ID 1DKZ / Resolution: 1.9 Å
• Authors: Zahn, M. / Straeter, N.
• Citation: Journal: To be Published; Title: Structural studies of DnaK in complex with proline rich antimicrobial peptides reveal two different peptide binding modes; Authors: Zahn, M. / Straeter, N.

History

Deposition	May 3, 2012	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Apr 24, 2013	Provider: repository / Type: Initial release
Revision 1.1	Nov 15, 2017	Group: Refinement description / Category: software / Item: _software.name
Revision 1.2	Feb 28, 2024	Group: Data collection / Database references Category: chem_comp_atom / chem_comp_bond / database_2 / struct_ref_seq_dif Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_ref_seq_dif.details

Assembly

Deposited unit

A: Chaperone protein DnaK

B: Proline rich bombesin-related protein

Summary:

• 25.9 kDa, 2 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	25,861	2
Polymers	25,861	2
Non-polymers	0	0
Water	1,441	80

1

A: Chaperone protein DnaK

B: Proline rich bombesin-related protein

A: Chaperone protein DnaK

B: Proline rich bombesin-related protein

A: Chaperone protein DnaK

B: Proline rich bombesin-related protein

A: Chaperone protein DnaK

B: Proline rich bombesin-related protein

Summary:

• defined by software
• 103 kDa, 8 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	103,445	8
Polymers	103,445	8
Non-polymers	0	0
Water	144	8

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1
crystal symmetry operation	2_675	-x+1,-y+2,z	1
crystal symmetry operation	3_657	-x+1,y,-z+2	1
crystal symmetry operation	4_577	x,-y+2,-z+2	1

Calculated values:

Buried area	16620 Å2
ΔGint	-108 kcal/mol
Surface area	39810 Å2
Method	PISA

2

A: Chaperone protein DnaK

B: Proline rich bombesin-related protein

A: Chaperone protein DnaK

B: Proline rich bombesin-related protein

Summary:

• defined by software
• 51.7 kDa, 4 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	51,722	4
Polymers	51,722	4
Non-polymers	0	0
Water	72	4

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1
crystal symmetry operation	3_657	-x+1,y,-z+2	1

Calculated values:

Buried area	6310 Å2
ΔGint	-41 kcal/mol
Surface area	21900 Å2
Method	PISA

3

A: Chaperone protein DnaK

B: Proline rich bombesin-related protein

A: Chaperone protein DnaK

B: Proline rich bombesin-related protein

Summary:

• defined by software
• 51.7 kDa, 4 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	51,722	4
Polymers	51,722	4
Non-polymers	0	0
Water	72	4

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1
crystal symmetry operation	4_577	x,-y+2,-z+2	1

Calculated values:

Buried area	5140 Å2
ΔGint	-36 kcal/mol
Surface area	23070 Å2
Method	PISA

4

Summary:

• Idetical with deposited unit
• defined by author&software

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1

Calculated values:

Buried area	1620 Å2
ΔGint	-12 kcal/mol
Surface area	12480 Å2
Method	PISA

Unit cell

Length a, b, c (Å)	96.645, 115.047, 39.366
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	23
Space group name H-M	I222

Components

#1: Protein

A Chaperone protein DnaK / HSP70 / Heat shock 70 kDa protein / Heat shock protein 70

Details:

Mass: 23820.777 Da / Num. of mol.: 1 / Fragment: UNP residues 389-607
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Strain: K12 / Gene: b0014, dnaK, groP, grpF, JW0013, seg / Production host: Escherichia coli (E. coli) / References: UniProt: P0A6Y8

#2: Protein/peptide

B Proline rich bombesin-related protein

Details:

Mass: 2040.453 Da / Num. of mol.: 1 / Fragment: UNP residues 46-60 / Source method: obtained synthetically
Details: This sequence occurs naturally in Bombina maxima. C-terminus is amidated.
Source: (synth.) Bombina maxima (large-webbed bell toad) / References: UniProt: Q8QFP2

#3: Water

ChemComp-HOH / water

Details:

Mass: 18.015 Da / Num. of mol.: 80 / Source method: isolated from a natural source / Formula: H₂O

Experimental details

Experiment

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

Sample preparation

• Crystal: Density Matthews: 2.11 ų/Da / Density % sol: 41.64 %
• Crystal grow: Temperature: 292 K / Method: vapor diffusion, hanging drop / pH: 8 ; Details: 3.0 M ammonium sulfate, 0.1 M HEPES pH 8.0, VAPOR DIFFUSION, HANGING DROP, temperature 292K

Data collection

• Diffraction: Mean temperature: 100 K
• Diffraction source: Source: SYNCHROTRON / Site: BESSY / Beamline: 14.2 / Wavelength: 0.91841 Å
• Detector: Type: MARMOSAIC 225 mm CCD / Detector: CCD / Date: Oct 19, 2010
• Radiation: Monochromator: Si - 111 / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
• Radiation wavelength: Wavelength: 0.91841 Å / Relative weight: 1
• Reflection: Resolution: 1.9→25 Å / Num. obs: 17741 / % possible obs: 99.8 % / R_merge(I) obs: 0.062
• Reflection shell: Resolution: 1.9→2 Å / Redundancy: 5.3 % / R_merge(I) obs: 0.558 / % possible all: 100

Processing

Software

Name	Version	Classification
MAR345		data collection
REFMAC	with PDBID 1DKZ	refinement
XDS		data reduction
SCALA		data scaling
REFMAC	with PDBID 1DKZ	phasing

Refinement

Method to determine structure: PDB ID 1DKZ / Resolution: 1.9→25 Å / Cor.coef. F_o:F_c: 0.948 / Cor.coef. F_o:F_c free: 0.916 / SU B: 9.73 / SU ML: 0.145 / Cross valid method: THROUGHOUT / ESU R: 0.187 / ESU R Free: 0.177 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN USED IF PRESENT IN THE INPUT

	Rfactor	Num. reflection	% reflection	Selection details
Rfree	0.27582	901	5.1 %	RANDOM
Rwork	0.21776	-	-	-
obs	0.22064	16807	99.57 %	-

• 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: 35.847 Ų

	Baniso -1	Baniso -2	Baniso -3
1-	-0.88 Å2	0 Å2	0 Å2
2-	-	-2.14 Å2	0 Å2
3-	-	-	3.02 Å2

Refinement step

Cycle: LAST / Resolution: 1.9→25 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	1749	0	0	80	1829

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.017	0.02	1795
X-RAY DIFFRACTION	r_bond_other_d
X-RAY DIFFRACTION	r_angle_refined_deg	1.931	1.966	2429
X-RAY DIFFRACTION	r_angle_other_deg
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	5.227	5	235
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	42.028	26.463	82
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	16.106	15	338
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	16.033	15	8
X-RAY DIFFRACTION	r_chiral_restr	0.129	0.2	283
X-RAY DIFFRACTION	r_gen_planes_refined	0.01	0.021	1335
X-RAY DIFFRACTION	r_gen_planes_other
X-RAY DIFFRACTION	r_nbd_refined
X-RAY DIFFRACTION	r_nbd_other
X-RAY DIFFRACTION	r_nbtor_refined
X-RAY DIFFRACTION	r_nbtor_other
X-RAY DIFFRACTION	r_xyhbond_nbd_refined
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
X-RAY DIFFRACTION	r_symmetry_vdw_other
X-RAY DIFFRACTION	r_symmetry_hbond_refined
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
X-RAY DIFFRACTION	r_mcbond_other
X-RAY DIFFRACTION	r_mcangle_it
X-RAY DIFFRACTION	r_scbond_it
X-RAY DIFFRACTION	r_scangle_it
X-RAY DIFFRACTION	r_rigid_bond_restr
X-RAY DIFFRACTION	r_sphericity_free
X-RAY DIFFRACTION	r_sphericity_bonded

LS refinement shell

Resolution: 1.9→1.949 Å / Total num. of bins used: 20

	Rfactor	Num. reflection	% reflection
Rfree	0.383	55	-
Rwork	0.272	1136	-
obs	-	-	99.92 %

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	0.6729	1.5744	0.9985	4.004	1.8062	2.4099	-0.0039	0.0424	-0.0111	-0.0035	0.0599	-0.0227	0.0523	0.0763	-0.0559	0.1502	0.0189	-0.0059	0.186	-0.0079	0.1382	62.5408	103.7951	38.1547
2	0.9589	-0.0549	0.5582	2.4311	-0.3189	1.1538	0.0533	0.0803	-0.2467	-0.2343	0.0744	-0.1019	0.107	0.0036	-0.1277	0.1538	0.0321	-0.0097	0.1397	-0.0041	0.1227	63.3594	90.5099	33.9227
3	2.3472	0.5126	-0.1734	5.4959	-1.1431	0.8878	-0.0905	0.2373	0.0056	-0.042	0.0325	0.0366	-0.0826	0.1262	0.058	0.1634	0.028	0.0172	0.1474	-0.0274	0.0505	61.3453	101.1881	30.3181
4	3.6556	2.8366	0.6523	11.3781	-1.1225	2.3661	0.2292	-0.0191	-0.2265	0.048	-0.1052	-0.4346	0.3095	0.0573	-0.124	0.1396	-0.0122	-0.0935	0.0862	-0.0163	0.1698	42.8047	74.7923	29.5546
5	5.003	-1.6968	0.4264	3.8476	-3.3948	4.9932	0.2401	0.1685	-0.169	-0.2762	0.0904	0.2326	0.3009	-0.0659	-0.3306	0.1583	-0.0766	-0.1393	0.1134	0.0183	0.1798	37.8018	77.1077	27.5935

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Auth asym-ID	Auth seq-ID
1	X-RAY DIFFRACTION	1	A	389 - 423
2	X-RAY DIFFRACTION	2	A	424 - 488
3	X-RAY DIFFRACTION	3	A	489 - 529
4	X-RAY DIFFRACTION	4	A	530 - 564
5	X-RAY DIFFRACTION	5	A	565 - 604