PDB-2ved: crystal structure of the chimerical mutant CapABK55M protein

Basic information

• Entry: Database: PDB / ID: 2ved
• Title: crystal structure of the chimerical mutant CapABK55M protein
• Components: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASEBiological membrane
• Keywords: TRANSFERASE / CO-POLYMERASE / ACTIVATION MECHANISM / CHIMERA / BACTERIAL TYROSINE-KINASE / EXOPOLYSACCHARIDE SYNTHESIS

Function / homology

Function:

carbohydrate:proton symporter activity / polysaccharide transport / extracellular polysaccharide biosynthetic process / non-specific protein-tyrosine kinase / non-membrane spanning protein tyrosine kinase activity / kinase activity / phosphorylation / nucleotide binding / ATP binding / plasma membrane

Domain/homology:

Polysaccharide export protein MPA1-like / Exopolysaccharide synthesis protein / CobQ/CobB/MinD/ParA nucleotide binding domain / Polysaccharide chain length determinant N-terminal domain / Chain length determinant protein / AAA domain / AAA domain / P-loop containing nucleotide triphosphate hydrolases / P-loop containing nucleoside triphosphate hydrolase / Rossmann fold / 3-Layer(aba) Sandwich / Alpha Beta

Component:

ADENOSINE-5'-DIPHOSPHATE / non-specific protein-tyrosine kinase / Capsular biosynthesis protein

• Biological species: STAPHYLOCOCCUS AUREUS (bacteria)
• Method: X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.6 Å
• Authors: Olivares-Illana, V. / Meyer, P. / Gueguen-Chaignon, V. / Soulat, D. / Deustcher, J. / Cozzone, A.J. / Morera, S. / Grangeasse, C. / Nessler, S.
• Citation: Journal: Plos Biol. / Year: 2008; Title: Structural Basis for the Regulation Mechanism of the Tyrosine Kinase Capb from Staphylococcus Aureus.; Authors: Olivares-Illana, V. / Meyer, P. / Bechet, E. / Gueguen-Chaignon, V. / Soulat, D. / Lazereg-Riquier, S. / Mijakovic, I. / Deutscher, J. / Cozzone, A.J. / Laprevote, O. / Morera, S. / Grangeasse, C. / Nessler, S.

History

Deposition	Oct 19, 2007	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Jul 1, 2008	Provider: repository / Type: Initial release
Revision 1.1	May 8, 2011	Group: Version format compliance
Revision 1.2	Jul 13, 2011	Group: Version format compliance
Revision 1.3	Dec 13, 2023	Group: Data collection / Database references / Derived calculations / Other / Refinement description Category: chem_comp_atom / chem_comp_bond / database_2 / pdbx_database_status / pdbx_initial_refinement_model / pdbx_struct_conn_angle / struct_conn Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_database_status.status_code_sf / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _struct_conn.pdbx_dist_value / _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_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_conn.ptnr2_label_seq_id

Assembly

Deposited unit

A: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

B: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

hetero molecules

Summary:

• 60.9 kDa, 2 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	60,905	6
Polymers	60,002	2
Non-polymers	903	4
Water	2,018	112

1

A: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

B: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

hetero molecules

A: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

B: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

hetero molecules

A: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

B: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

hetero molecules

A: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

B: MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE

hetero molecules

Summary:

• defined by software
• 244 kDa, 8 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	243,618	24
Polymers	240,006	8
Non-polymers	3,612	16
Water	144	8

Symmetry operations:

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

Calculated values:

Buried area	15940 Å2
ΔGint	-47.9 kcal/mol
Surface area	91770 Å2
Method	PQS

Unit cell

Length a, b, c (Å)	163.200, 163.200, 57.200
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	79
Space group name H-M	I4

Components

#1: Protein

A B MEMBRANE PROTEIN CAPA1, PROTEIN TYROSINE KINASE / Biological membrane / CHIMERICAL PROTEIN CAPAB / CAP5A

Details:

Mass: 30000.756 Da / Num. of mol.: 2 / Fragment: THE LAST 29AA FROM CAPA1, RESIDUES 197-222, / Mutation: YES
Source method: isolated from a genetically manipulated source
Details: BOUND ADP AND MG / Source: (gene. exp.) STAPHYLOCOCCUS AUREUS (bacteria) / Gene: CAPA1, CAPB2 / Plasmid: PQE30 / Production host: ESCHERICHIA COLI (E. coli) / Strain (production host): BL21 / References: UniProt: A8YPQ6, UniProt: A8YPQ5

#2: Chemical

A-2227 B-2229 ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE / Adenosine diphosphate

Details:

Mass: 427.201 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₁₀H₁₅N₅O₁₀P₂ / Comment: ADP, energy-carrying molecule*YM

#3: Chemical

A-2228 B-2230 ChemComp-MG / MAGNESIUM ION

Details:

Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg

#4: Water

ChemComp-HOH / water / Water

Details:

Mass: 18.015 Da / Num. of mol.: 112 / Source method: isolated from a natural source / Formula: H₂O

• Compound details: ENGINEERED RESIDUE IN CHAIN A, LYS 55 TO MET ENGINEERED RESIDUE IN CHAIN B, LYS 55 TO MET
• Sequence details: THIS IS A CHIMERICAL PROTEIN COMPOSED OF THE 29AA OF THE C-TERMINAL DOMAIN OF CAPA1 (RESIDUE NAME FROM 197 TO 222) IN FUSION WITH CAPB2 PROTEIN. THERE ARE TWO CHIMERICAL PROTEINS IN THE ASYMMETRIC UNIT

Experimental details

Experiment

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

Sample preparation

• Crystal: Density Matthews: 3.32 ų/Da / Density % sol: 62.71 % / Description: NONE
• Crystal grow: pH: 7.5 / Details: 20% PEG1000 100MM HEPES PH7.5 200MM GLYCINE

Data collection

• Diffraction: Mean temperature: 100 K
• Diffraction source: Source: SYNCHROTRON / Site: ESRF / Beamline: ID29 / Wavelength: 0.9
• Detector: Type: ADSC QUANTUM 315r / Detector: CCD / Date: May 19, 2007
• Radiation: Monochromator: SI / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
• Radiation wavelength: Wavelength: 0.9 Å / Relative weight: 1
• Reflection: Resolution: 2.6→20 Å / Num. obs: 23263 / % possible obs: 99.4 % / Observed criterion σ(I): 1 / Redundancy: 4 % / R_merge(I) obs: 0.15 / Net I/σ(I): 8
• Reflection shell: Resolution: 2.6→2.7 Å / Redundancy: 3 % / R_merge(I) obs: 0.59 / Mean I/σ(I) obs: 1.9 / % possible all: 96.4

Processing

Software

Name	Version	Classification
REFMAC	5.2.0019	refinement
MOSFLM		data reduction
SCALA		data scaling
PHASER		phasing

Refinement

Method to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 1ION

1ion

Resolution: 2.6→19.79 Å / Cor.coef. F_o:F_c: 0.922 / Cor.coef. F_o:F_c free: 0.854 / SU B: 11.616 / SU ML: 0.241 / Cross valid method: THROUGHOUT / ESU R: 0.457 / ESU R Free: 0.307 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS

	Rfactor	Num. reflection	% reflection	Selection details
Rfree	0.27	1164	5 %	RANDOM
Rwork	0.204	-	-	-
obs	0.208	22099	100 %	-

• Solvent computation: Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.4 Å / Solvent model: MASK

Displacement parameters

B_iso mean: 28.42 Ų

	Baniso -1	Baniso -2	Baniso -3
1-	-0.01 Å2	0 Å2	0 Å2
2-	-	-0.01 Å2	0 Å2
3-	-	-	0.03 Å2

Refinement step

Cycle: LAST / Resolution: 2.6→19.79 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	3923	0	56	112	4091

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.017	0.022	4053
X-RAY DIFFRACTION	r_bond_other_d
X-RAY DIFFRACTION	r_angle_refined_deg	1.87	1.991	5519
X-RAY DIFFRACTION	r_angle_other_deg
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	9.073	5	504
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	39.091	25.607	173
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	18.061	15	694
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	20.341	15	16
X-RAY DIFFRACTION	r_chiral_restr	0.134	0.2	639
X-RAY DIFFRACTION	r_gen_planes_refined	0.006	0.02	3014
X-RAY DIFFRACTION	r_gen_planes_other
X-RAY DIFFRACTION	r_nbd_refined	0.238	0.2	1968
X-RAY DIFFRACTION	r_nbd_other
X-RAY DIFFRACTION	r_nbtor_refined	0.321	0.2	2850
X-RAY DIFFRACTION	r_nbtor_other
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.161	0.2	195
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	0.198	0.2	45
X-RAY DIFFRACTION	r_symmetry_vdw_other
X-RAY DIFFRACTION	r_symmetry_hbond_refined	0.19	0.2	11
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	0.907	1.5	2584
X-RAY DIFFRACTION	r_mcbond_other
X-RAY DIFFRACTION	r_mcangle_it	1.556	2	4117
X-RAY DIFFRACTION	r_mcangle_other
X-RAY DIFFRACTION	r_scbond_it	2.021	3	1661
X-RAY DIFFRACTION	r_scbond_other
X-RAY DIFFRACTION	r_scangle_it	3.283	4.5	1402
X-RAY DIFFRACTION	r_scangle_other
X-RAY DIFFRACTION	r_long_range_B_refined
X-RAY DIFFRACTION	r_long_range_B_other
X-RAY DIFFRACTION	r_rigid_bond_restr
X-RAY DIFFRACTION	r_sphericity_free
X-RAY DIFFRACTION	r_sphericity_bonded

LS refinement shell

Resolution: 2.6→2.67 Å / Total num. of bins used: 20

	Rfactor	Num. reflection	% reflection
Rfree	0.354	80	-
Rwork	0.29	1516	-
obs	-	-	100 %