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- PDB-5fb4: Crystal structure of the bacteriophage phi29 tail knob protein gp... -

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Basic information

Entry
Database: PDB / ID: 5fb4
TitleCrystal structure of the bacteriophage phi29 tail knob protein gp9 truncation variant
ComponentsDistal tube protein
KeywordsVIRAL PROTEIN / bacteriophage phi29 / tail knob
Function / homology
Function and homology information


virus tail, tube / symbiont genome ejection through host cell envelope, short tail mechanism / virus tail / symbiont genome entry into host cell via pore formation in plasma membrane
Similarity search - Function
: / Bacillus phage phi29, Tail knob protein gp9, C-terminal domain / Distal tube protein, N-terminal / Caudoviral major tail protein N-terminus
Similarity search - Domain/homology
DI(HYDROXYETHYL)ETHER / Tail knob protein gp9
Similarity search - Component
Biological speciesBacillus phage phi29 (virus)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.039 Å
AuthorsXu, J.W. / Gui, M. / Wang, D.H. / Xiang, Y.
Funding support China, 4items
OrganizationGrant numberCountry
973 program2015CB13910102 China
the National Natural Science Foundation of China31470721 China
Science Foundation of China81550001 China
the Junior Thousand Talents Program of China201311770418 China
CitationJournal: Nature / Year: 2016
Title: The bacteriophage ϕ29 tail possesses a pore-forming loop for cell membrane penetration.
Authors: Jingwei Xu / Miao Gui / Dianhong Wang / Ye Xiang /
Abstract: Most bacteriophages are tailed bacteriophages with an isometric or a prolate head attached to a long contractile, long non-contractile, or short non-contractile tail. The tail is a complex machine ...Most bacteriophages are tailed bacteriophages with an isometric or a prolate head attached to a long contractile, long non-contractile, or short non-contractile tail. The tail is a complex machine that plays a central role in host cell recognition and attachment, cell wall and membrane penetration, and viral genome ejection. The mechanisms involved in the penetration of the inner host cell membrane by bacteriophage tails are not well understood. Here we describe structural and functional studies of the bacteriophage ϕ29 tail knob protein gene product 9 (gp9). The 2.0 Å crystal structure of gp9 shows that six gp9 molecules form a hexameric tube structure with six flexible hydrophobic loops blocking one end of the tube before DNA ejection. Sequence and structural analyses suggest that the loops in the tube could be membrane active. Further biochemical assays and electron microscopy structural analyses show that the six hydrophobic loops in the tube exit upon DNA ejection and form a channel that spans the lipid bilayer of the membrane and allows the release of the bacteriophage genomic DNA, suggesting that cell membrane penetration involves a pore-forming mechanism similar to that of certain non-enveloped eukaryotic viruses. A search of other phage tail proteins identified similar hydrophobic loops, which indicates that a common mechanism might be used for membrane penetration by prokaryotic viruses. These findings suggest that although prokaryotic and eukaryotic viruses use apparently very different mechanisms for infection, they have evolved similar mechanisms for breaching the cell membrane.
History
DepositionDec 14, 2015Deposition site: RCSB / Processing site: PDBJ
Revision 1.0Jun 22, 2016Provider: repository / Type: Initial release
Revision 1.1Jun 29, 2016Group: Database references
Revision 1.2Nov 8, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / citation / database_2 / pdbx_initial_refinement_model / pdbx_struct_oper_list
Item: _citation.journal_id_CSD / _database_2.pdbx_DOI ..._citation.journal_id_CSD / _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_oper_list.symmetry_operation

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Distal tube protein
B: Distal tube protein
C: Distal tube protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)205,5216
Polymers205,2023
Non-polymers3183
Water23,6361312
1
A: Distal tube protein
B: Distal tube protein
C: Distal tube protein
hetero molecules

A: Distal tube protein
B: Distal tube protein
C: Distal tube protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)411,04112
Polymers410,4056
Non-polymers6376
Water1086
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_755-x+2,-y,z1
Buried area37170 Å2
ΔGint-144 kcal/mol
Surface area112500 Å2
MethodPISA
Unit cell
Length a, b, c (Å)94.600, 135.161, 313.431
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number23
Space group name H-MI222

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Components

#1: Protein Distal tube protein / Gene product 9 / gp9


Mass: 68400.758 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacillus phage phi29 (virus) / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 / References: UniProt: P04331
#2: Chemical ChemComp-PEG / DI(HYDROXYETHYL)ETHER


Mass: 106.120 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C4H10O3
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 1312 / Source method: isolated from a natural source / Formula: H2O
Sequence detailsTHE EXPERIMENTAL INFO OF UNIPROT (P04331, TUB9_BPPH2) REPORTS CONFLICT AT THESE POSITIONS 21 AND ...THE EXPERIMENTAL INFO OF UNIPROT (P04331, TUB9_BPPH2) REPORTS CONFLICT AT THESE POSITIONS 21 AND 141 (in Ref. 2; ACE96032).

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.44 Å3/Da / Density % sol: 49.62 %
Crystal growTemperature: 293.15 K / Method: vapor diffusion, hanging drop / pH: 7.5
Details: 0.94% v/v ethanol, 10% PEG-400, HEPES buffer pH 7.5, magnesium chloride

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Data collection

DiffractionMean temperature: 160 K
Diffraction sourceSource: SYNCHROTRON / Site: SSRF / Beamline: BL17U / Wavelength: 0.978 Å
DetectorType: RAYONIX MX-225 / Detector: CCD / Date: Oct 3, 2014
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.978 Å / Relative weight: 1
ReflectionResolution: 2.039→43.32 Å / Num. obs: 126814 / % possible obs: 99.29 % / Redundancy: 6.1 % / Net I/σ(I): 17.66
Reflection shellRedundancy: 5.9 % / Mean I/σ(I) obs: 9.2 / % possible all: 95.11

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Processing

Software
NameVersionClassification
PHENIX1.9_1692refinement
HKL-2000data collection
HKL-2000data scaling
SHARPphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5FEI

5fei


Resolution: 2.039→43.317 Å / SU ML: 0.16 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 17.99 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.1832 6371 5.02 %
Rwork0.1531 --
obs0.1547 126809 99.29 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 2.039→43.317 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms12499 0 21 1312 13832
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00812816
X-RAY DIFFRACTIONf_angle_d1.05317336
X-RAY DIFFRACTIONf_dihedral_angle_d11.794709
X-RAY DIFFRACTIONf_chiral_restr0.0481843
X-RAY DIFFRACTIONf_plane_restr0.0052240
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.0392-2.06240.20871980.17053389X-RAY DIFFRACTION85
2.0624-2.08670.19982080.1643986X-RAY DIFFRACTION100
2.0867-2.11210.17932030.15844045X-RAY DIFFRACTION100
2.1121-2.13890.21082360.15613954X-RAY DIFFRACTION100
2.1389-2.1670.19612290.15373986X-RAY DIFFRACTION100
2.167-2.19670.19182060.15684001X-RAY DIFFRACTION100
2.1967-2.22810.19781850.15754039X-RAY DIFFRACTION100
2.2281-2.26130.18362050.14984034X-RAY DIFFRACTION100
2.2613-2.29660.18412010.15144026X-RAY DIFFRACTION100
2.2966-2.33430.20482020.15564004X-RAY DIFFRACTION100
2.3343-2.37450.19422230.1644011X-RAY DIFFRACTION100
2.3745-2.41770.21652280.1634006X-RAY DIFFRACTION100
2.4177-2.46420.20092120.1684017X-RAY DIFFRACTION100
2.4642-2.51450.21981910.16714018X-RAY DIFFRACTION100
2.5145-2.56920.22731870.16734072X-RAY DIFFRACTION100
2.5692-2.62890.2282280.16674001X-RAY DIFFRACTION100
2.6289-2.69470.20122160.16374021X-RAY DIFFRACTION100
2.6947-2.76750.21931910.1664066X-RAY DIFFRACTION100
2.7675-2.84890.18452130.16394030X-RAY DIFFRACTION100
2.8489-2.94090.19991990.16264043X-RAY DIFFRACTION100
2.9409-3.0460.20352130.16394050X-RAY DIFFRACTION100
3.046-3.16790.19082070.15854063X-RAY DIFFRACTION100
3.1679-3.3120.18152100.14814038X-RAY DIFFRACTION100
3.312-3.48660.16572240.14864061X-RAY DIFFRACTION100
3.4866-3.70490.14832280.14154036X-RAY DIFFRACTION100
3.7049-3.99080.16542320.13334056X-RAY DIFFRACTION100
3.9908-4.3920.15932280.13044058X-RAY DIFFRACTION100
4.392-5.02670.15232180.12814129X-RAY DIFFRACTION100
5.0267-6.330.17362190.15824136X-RAY DIFFRACTION100
6.33-43.32720.17282310.17174062X-RAY DIFFRACTION95
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°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)
10.4241-0.03130.16050.31490.29571.9209-0.0572-0.14160.05130.1630.0364-0.025-0.06440.10470.00440.20210.0084-0.0140.1282-0.02290.186994.100630.1694100.8689
21.9801-0.2591-0.26451.3621-0.17031.42180.02730.2560.1328-0.14710.0060.021-0.10230.04940.0060.1625-0.0183-0.01220.19870.04680.165896.181433.50418.9752
30.2554-0.0255-0.1230.1323-0.16441.4555-0.01850.02630.03790.01080.0140.0022-0.03970.02650.00690.1128-0.0109-0.01570.0779-0.00110.164195.749527.882561.1693
40.7367-0.20310.68460.5916-0.26381.6312-0.1143-0.30930.07610.32620.0419-0.1063-0.1407-0.1775-0.01590.25130.0198-0.0720.215-0.03640.2321121.102215.9926100.8909
51.1734-0.0543-0.14412.0310.09251.45240.02720.1430.0188-0.2009-0.0007-0.08620.01320.1020.01330.1097-0.00910.01170.20950.02220.1527124.238215.150518.798
60.1798-0.0307-0.080.24680.10921.6294-0.00860.03240.02320.0173-0.0063-0.03830.0132-0.020.01580.0936-0.0254-0.01930.11330.01040.1828119.157412.968660.215
70.4887-0.04180.1550.6647-0.16660.47730.0734-0.3148-0.03270.5196-0.2176-0.3351-0.20360.5040.03510.3784-0.0465-0.12380.38380.07360.3005125.6408-16.6797107.9861
80.5411-0.03570.06561.05380.04350.53920.055-0.43920.01310.9066-0.23680.2408-0.375-0.0247-0.00190.6824-0.0549-0.10340.39370.0410.3825120.8947-13.7519116.4817
90.36850.1247-0.26220.7268-0.68840.62160.0402-0.0755-0.02370.2343-0.0535-0.0549-0.21140.16-0.01070.1645-0.0095-0.04140.1052-0.00190.1607118.9008-13.346788.5369
101.70560.4911-0.01491.58020.16061.4476-0.05130.2317-0.0224-0.21580.0142-0.10470.0760.07350.01830.15510.01550.03970.1915-0.00060.1704122.8479-18.302718.9188
110.21920.0425-0.14250.19680.09541.8364-0.01040.0231-0.0105-0.0037-0.0059-0.0544-0.02990.03390.0210.06830.0204-0.00290.11050.00620.1804118.0366-14.621360.1963
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection details
1X-RAY DIFFRACTION1chain A and (resid 2 through 165 )
2X-RAY DIFFRACTION2chain A and (resid 166 through 293 )
3X-RAY DIFFRACTION3chain A and (resid 294 through 599 )
4X-RAY DIFFRACTION4chain B and (resid 2 through 165 )
5X-RAY DIFFRACTION5chain B and (resid 166 through 293 )
6X-RAY DIFFRACTION6chain B and (resid 294 through 599 )
7X-RAY DIFFRACTION7chain C and (resid 2 through 52 )
8X-RAY DIFFRACTION8chain C and (resid 53 through 89 )
9X-RAY DIFFRACTION9chain C and (resid 90 through 165 )
10X-RAY DIFFRACTION10chain C and (resid 166 through 293 )
11X-RAY DIFFRACTION11chain C and (resid 294 through 599 )

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