PDB-4qf0: Structure of a 16 nm protein cage designed by fusing symmetric ol...

Basic information

• Entry: Database: PDB / ID: 4qf0
• Title: Structure of a 16 nm protein cage designed by fusing symmetric oligomeric domains, quadruple mutant, P21212 form
• Components: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera
• Keywords: OXIDOREDUCTASE / VIRAL PROTEIN / protein design / bionanotechnology / protein assembly / symmetry / biomaterials

Function / homology

Function:

Assembly of Viral Components at the Budding Site / Influenza Infection / Fusion of the Influenza Virion to the Host Cell Endosome / Release / Budding / Packaging of Eight RNA Segments / Uncoating of the Influenza Virion / Entry of Influenza Virion into Host Cell via Endocytosis / Viral RNP Complexes in the Host Cell Nucleus / NEP/NS2 Interacts with the Cellular Export Machinery / Viral mRNA Translation / antibiotic biosynthetic process / Oxidoreductases; Acting on a peroxide as acceptor; Peroxidases / viral budding from plasma membrane / peroxidase activity / structural constituent of virion / host cell nucleus / virion membrane / RNA binding / extracellular region / plasma membrane

Domain/homology:

Matrix protein 1 / Influenza matrix M1, N-terminal / Influenza matrix M1, C-terminal / Influenza matrix M1, N-terminal subdomain 1 / Influenza matrix M1, N-terminal subdomain 2 / Influenza virus matrix protein M1 / Influenza Matrix protein (M1) / Influenza Matrix protein (M1) C-terminal domain / Influenza Matrix protein (M1) C-terminal domain / Epoxide hydrolase-like / alpha/beta hydrolase fold / Alpha/beta hydrolase fold-1 / Alpha/Beta hydrolase fold

Component:

Matrix protein 1 / Non-haem bromoperoxidase BPO-A2

• Biological species: Streptomyces aureofaciens (bacteria); Influenza A virus
• Method: X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 6.494 Å
• Authors: Lai, Y.-T. / Yeates, T.O.
• Citation: Journal: To be Published; Title: Structural transition of a protein nanocage as a function of pH and salt concentration.; Authors: Lai, Y.-T. / Yeates, T.O.

History

Deposition	May 19, 2014	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	May 20, 2015	Provider: repository / Type: Initial release
Revision 1.1	Jul 26, 2017	Group: Refinement description / Source and taxonomy / Category: entity_src_gen / software
Revision 1.2	Feb 28, 2024	Group: Data collection / Database references / Refinement description Category: chem_comp_atom / chem_comp_bond / database_2 / struct_ncs_dom_lim / struct_ref_seq_dif Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id / _struct_ncs_dom_lim.beg_label_comp_id / _struct_ncs_dom_lim.beg_label_seq_id / _struct_ncs_dom_lim.end_auth_comp_id / _struct_ncs_dom_lim.end_label_asym_id / _struct_ncs_dom_lim.end_label_comp_id / _struct_ncs_dom_lim.end_label_seq_id / _struct_ref_seq_dif.details

Assembly

Deposited unit

A: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

B: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

C: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

D: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

E: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

F: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

Summary:

• 301 kDa, 6 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	301,136	6
Polymers	301,136	6
Non-polymers	0	0
Water	0	0

1

A: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

B: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

C: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

D: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

E: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

F: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

A: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

B: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

C: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

D: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

E: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

F: Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera

Summary:

• defined by author&software
• 602 kDa, 12 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	602,271	12
Polymers	602,271	12
Non-polymers	0	0
Water	0

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1
crystal symmetry operation	2_465	-x-1,-y+1,z	1

Calculated values:

Buried area	33870 Å2
ΔGint	-89 kcal/mol
Surface area	194430 Å2
Method	PISA

Unit cell

Length a, b, c (Å)	175.500, 147.730, 167.630
Angle α, β, γ (deg.)	90.000, 90.000, 90.000
Int Tables number	18
Space group name H-M	P21212

Noncrystallographic symmetry (NCS)

NCS domain:

ID	Ens-ID	Details
1	1	A
2	1	B
1	2	A
2	2	C
1	3	A
2	3	D
1	4	A
2	4	E
1	5	A
2	5	F
1	6	B
2	6	C
1	7	B
2	7	D
1	8	B
2	8	E
1	9	B
2	9	F
1	10	C
2	10	D
1	11	C
2	11	E
1	12	C
2	12	F
1	13	D
2	13	E
1	14	D
2	14	F
1	15	E
2	15	F

NCS domain segments:

Component-ID: _ / Beg auth comp-ID: PRO / Beg label comp-ID: PRO / End auth comp-ID: GLN / End label comp-ID: GLN / Refine code: _ / Auth seq-ID: 1 - 441 / Label seq-ID: 2 - 442

Dom-ID	Ens-ID	Auth asym-ID	Label asym-ID
1	1	A	A
2	1	B	B
1	2	A	A
2	2	C	C
1	3	A	A
2	3	D	D
1	4	A	A
2	4	E	E
1	5	A	A
2	5	F	F
1	6	B	B
2	6	C	C
1	7	B	B
2	7	D	D
1	8	B	B
2	8	E	E
1	9	B	B
2	9	F	F
1	10	C	C
2	10	D	D
1	11	C	C
2	11	E	E
1	12	C	C
2	12	F	F
1	13	D	D
2	13	E	E
1	14	D	D
2	14	F	F
1	15	E	E
2	15	F	F

NCS ensembles :

ID
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15

Components

#1: Protein

A B C D E F
Non-haem bromoperoxidase BPO-A2, Matrix protein 1 chimera / Designed 16nm tetrahedral protein cage / BPO2 / Bromide peroxidase / M1

Details:

Mass: 50189.266 Da / Num. of mol.: 6 / Fragment: SEE REMARK 999 / Mutation: K118A, L279Q, Q24T, Y51A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Streptomyces aureofaciens (bacteria), (gene. exp.) Influenza A virus
Gene: bpoA2, M / Plasmid: pET22b / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3)
References: UniProt: P29715, UniProt: P03485, Oxidoreductases; Acting on a peroxide as acceptor; Peroxidases

• Sequence details: THE DESIGNED PROTEIN CONSTRUCT IS A CHIMERA COMPRISING BPO2 (UNP P29715) AND RESIDUES 2-164 (UNP P03485) OF M1, SEPARATED BY A LINKER.

Experimental details

Experiment

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

Sample preparation

• Crystal: Density Matthews: 3.61 ų/Da / Density % sol: 65.91 %
• Crystal grow: Temperature: 298 K / Method: vapor diffusion, hanging drop / pH: 7 ; Details: 0.1 M Tris, pH 7.0, 10% PEG8000, 0.2 M magnesium chloride, 3% trehalose, VAPOR DIFFUSION, HANGING DROP, temperature 298K

Data collection

• Diffraction: Mean temperature: 100 K
• Diffraction source: Source: SYNCHROTRON / Site: APS / Beamline: 24-ID-C / Wavelength: 0.9792 Å
• Detector: Type: DECTRIS PILATUS 6M-F / Detector: PIXEL / Date: Dec 8, 2013
• Radiation: Monochromator: Cryo-Cooled double crystal Si(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
• Radiation wavelength: Wavelength: 0.9792 Å / Relative weight: 1
• Reflection: Resolution: 6.49→93.71 Å / Num. obs: 9001 / % possible obs: 99.6 % / Observed criterion σ(I): -3 / B_iso Wilson estimate: 410.357 Ų / R_merge(I) obs: 0.086 / Χ²: 1.017 / Net I/σ(I): 14.16

Reflection shell

Resolution (Å)	Rmerge(I) obs	Mean I/σ(I) obs	Num. measured obs	Num. unique obs	Diffraction-ID	% possible all
6.49-6.66	0.819	2.34	3734	613	1	96.1
6.66-6.84	0.74	2.76	4304	628	1	99.8
6.84-7.04	0.558	3.52	4166	630	1	100
7.04-7.26	0.477	4.16	3965	596	1	100
7.26-7.5	0.378	5.43	3844	594	1	100
7.5-7.76	0.309	6.25	3473	568	1	100
7.76-8.05	0.229	7.9	3009	544	1	99.3
8.05-8.38	0.167	10.69	3167	531	1	100
8.38-8.76	0.12	15.55	3505	508	1	100
8.76-9.18	0.1	17	3293	492	1	100
9.18-9.68	0.084	20.6	3149	469	1	100
9.68-10.27	0.08	22.57	2813	436	1	100
10.27-10.98	0.066	23.74	2739	427	1	100
10.98-11.86	0.06	26.34	2507	394	1	100
11.86-12.99	0.055	26.89	2281	362	1	100
12.99-14.52	0.051	26.91	1908	326	1	100
14.52-16.77	0.046	25.71	1470	298	1	98.7
16.77-20.53	0.044	33.55	1668	256	1	100
20.53-29.04	0.04	35.6	1299	207	1	100
29.04-93.71	0.047	36.26	664	122	1	95.3

Phasing

• Phasing: Method: molecular replacement

Phasing MR

Model details: Phaser MODE: MR_AUTO

	Highest resolution	Lowest resolution
Rotation	6.49 Å	93.71 Å
Translation	6.49 Å	93.71 Å

Processing

Software

Name	Version	Classification	NB
XSCALE		data scaling
PHASER	2.5.6	phasing
REFMAC	5.8.0071	refinement
PDB_EXTRACT	3.14	data extraction

Refinement

Method to determine structure: MOLECULAR REPLACEMENT / Resolution: 6.494→93.71 Å / Cor.coef. F_o:F_c: 0.841 / Cor.coef. F_o:F_c free: 0.926 / WR_factor R_free: 0.3242 / WR_factor R_work: 0.271 / FOM work R set: 0.7439 / SU B: 275.849 / SU ML: 2.448 / SU R_free: 3.3017 / Cross valid method: THROUGHOUT / σ(F): 0 / ESU R Free: 3.302 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS

	Rfactor	Num. reflection	% reflection	Selection details
Rfree	0.3242	451	5 %	RANDOM
Rwork	0.271	-	-	-
obs	0.2739	8551	99.56 %	-

• 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 max: 340 Ų / B_iso mean: 309.413 Ų / B_iso min: 114.51 Ų

	Baniso -1	Baniso -2	Baniso -3
1-	26.11 Å2	0 Å2	-0 Å2
2-	-	-14.18 Å2	0 Å2
3-	-	-	-11.93 Å2

Refinement step

Cycle: LAST / Resolution: 6.494→93.71 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	20256	0	0	0	20256

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.007	0.019	20718
X-RAY DIFFRACTION	r_bond_other_d	0.003	0.02	19332
X-RAY DIFFRACTION	r_angle_refined_deg	1.007	1.953	28224
X-RAY DIFFRACTION	r_angle_other_deg	1.642	3	44298
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	4.82	5	2640
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	33.366	23.742	930
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	14.263	15	3150
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	13.41	15	132
X-RAY DIFFRACTION	r_chiral_restr	0.051	0.2	3210
X-RAY DIFFRACTION	r_gen_planes_refined	0.003	0.021	23892
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	4872
X-RAY DIFFRACTION	r_mcbond_it	8.534	30.963	10578
X-RAY DIFFRACTION	r_mcbond_other	8.535	30.963	10577
X-RAY DIFFRACTION	r_mcangle_it	14.527	46.415	13212

Refine LS restraints NCS

Refine-ID: X-RAY DIFFRACTION / Type: interatomic distance / Weight position: 0.05

Ens-ID	Dom-ID	Auth asym-ID	Number	Rms dev position (Å)
1	1	A	26604	0.01
1	2	B	26604	0.01
2	1	A	26582	0.02
2	2	C	26582	0.02
3	1	A	26613	0.01
3	2	D	26613	0.01
4	1	A	26610	0.01
4	2	E	26610	0.01
5	1	A	26609	0.02
5	2	F	26609	0.02
6	1	B	26595	0.02
6	2	C	26595	0.02
7	1	B	26619	0.01
7	2	D	26619	0.01
8	1	B	26612	0.01
8	2	E	26612	0.01
9	1	B	26616	0.01
9	2	F	26616	0.01
10	1	C	26594	0.02
10	2	D	26594	0.02
11	1	C	26596	0.02
11	2	E	26596	0.02
12	1	C	26591	0.02
12	2	F	26591	0.02
13	1	D	26618	0.01
13	2	E	26618	0.01
14	1	D	26615	0.01
14	2	F	26615	0.01
15	1	E	26626	0.01
15	2	F	26626	0.01

LS refinement shell

Resolution: 6.494→6.661 Å / Total num. of bins used: 20

	Rfactor	Num. reflection	% reflection
Rfree	0.413	31	-
Rwork	0.344	580	-
all	-	611	-
obs	-	-	96.07 %