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- PDB-4zuy: Structure of Tsi6 from Pseudomonas aeruginosa -

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Basic information

Entry
Database: PDB / ID: 4zuy
TitleStructure of Tsi6 from Pseudomonas aeruginosa
ComponentsTsi6
KeywordsPROTEIN BINDING / 4-helix bundle / bacterial type VI secretion immunity protein / Tse6 immunity protein
Function / homologyTsi6 / Tsi6 / protein secretion by the type VI secretion system / toxin sequestering activity / Immune protein Tsi6
Function and homology information
Biological speciesPseudomonas aeruginosa (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / SAD / Resolution: 1.952 Å
AuthorsWhitney, J.C. / Sawai, S. / Robinson, H. / Mougous, J.D.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI080609 United States
Defense Threat Reduction Agency (DTRA)HDTRA-1-13-014 United States
CitationJournal: Cell / Year: 2015
Title: An interbacterial NAD(P)(+) glycohydrolase toxin requires elongation factor Tu for delivery to target cells.
Authors: John C Whitney / Dennis Quentin / Shin Sawai / Michele LeRoux / Brittany N Harding / Hannah E Ledvina / Bao Q Tran / Howard Robinson / Young Ah Goo / David R Goodlett / Stefan Raunser / Joseph D Mougous /
Abstract: Type VI secretion (T6S) influences the composition of microbial communities by catalyzing the delivery of toxins between adjacent bacterial cells. Here, we demonstrate that a T6S integral membrane ...Type VI secretion (T6S) influences the composition of microbial communities by catalyzing the delivery of toxins between adjacent bacterial cells. Here, we demonstrate that a T6S integral membrane toxin from Pseudomonas aeruginosa, Tse6, acts on target cells by degrading the universally essential dinucleotides NAD(+) and NADP(+). Structural analyses of Tse6 show that it resembles mono-ADP-ribosyltransferase proteins, such as diphtheria toxin, with the exception of a unique loop that both excludes proteinaceous ADP-ribose acceptors and contributes to hydrolysis. We find that entry of Tse6 into target cells requires its binding to an essential housekeeping protein, translation elongation factor Tu (EF-Tu). These proteins participate in a larger assembly that additionally directs toxin export and provides chaperone activity. Visualization of this complex by electron microscopy defines the architecture of a toxin-loaded T6S apparatus and provides mechanistic insight into intercellular membrane protein delivery between bacteria.
History
DepositionMay 18, 2015Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 11, 2015Provider: repository / Type: Initial release
Revision 1.1Sep 20, 2017Group: Author supporting evidence / Derived calculations / Category: pdbx_audit_support / pdbx_struct_oper_list
Item: _pdbx_audit_support.funding_organization / _pdbx_struct_oper_list.symmetry_operation
Revision 1.2Apr 24, 2019Group: Author supporting evidence / Data collection / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.3Dec 11, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Tsi6
B: Tsi6
hetero molecules


Theoretical massNumber of molelcules
Total (without water)23,8946
Polymers23,7532
Non-polymers1424
Water1,76598
1
A: Tsi6
hetero molecules


Theoretical massNumber of molelcules
Total (without water)11,9834
Polymers11,8761
Non-polymers1063
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
2
B: Tsi6
hetero molecules


Theoretical massNumber of molelcules
Total (without water)11,9122
Polymers11,8761
Non-polymers351
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
3


  • Idetical with deposited unit
  • defined by software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area2280 Å2
ΔGint-47 kcal/mol
Surface area10210 Å2
MethodPISA
Unit cell
Length a, b, c (Å)54.511, 100.956, 93.098
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number20
Space group name H-MC2221

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Components

#1: Protein Tsi6


Mass: 11876.306 Da / Num. of mol.: 2 / Mutation: I37M
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas aeruginosa (strain ATCC 15692 / PAO1 / 1C / PRS 101 / LMG 12228) (bacteria)
Strain: ATCC 15692 / PAO1 / 1C / PRS 101 / LMG 12228 / Gene: PA0092 / Plasmid: pET29b / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 pLysS / References: UniProt: Q9I740
#2: Chemical
ChemComp-CL / CHLORIDE ION


Mass: 35.453 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Cl
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 98 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION

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Sample preparation

CrystalDensity Matthews: 2.72 Å3/Da / Density % sol: 54.74 %
Crystal growTemperature: 298 K / Method: vapor diffusion, hanging drop / Details: 0.1 M imidazole:HCl pH 8.0, 2.5 M NaCl

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: NSLS / Beamline: X29A / Wavelength: 0.979 Å
DetectorType: ADSC QUANTUM 315r / Detector: CCD / Date: Apr 8, 2014
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.979 Å / Relative weight: 1
ReflectionResolution: 1.95→47.98 Å / Num. obs: 19003 / % possible obs: 99.8 % / Redundancy: 15 % / Rmerge(I) obs: 0.097 / Net I/σ(I): 26
Reflection shellResolution: 1.95→2.02 Å / Rmerge(I) obs: 0.687

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Processing

Software
NameVersionClassification
PHENIX1.8.4_1496refinement
HKL-2000data reduction
HKL-2000data scaling
SHELXDEphasing
RefinementMethod to determine structure: SAD / Resolution: 1.952→47.966 Å / SU ML: 0.22 / Cross valid method: FREE R-VALUE / σ(F): 0 / Phase error: 24.29 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2356 1869 9.96 %
Rwork0.1918 --
obs0.196 18765 98.47 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 1.952→47.966 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1451 0 4 98 1553
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0071472
X-RAY DIFFRACTIONf_angle_d0.9331991
X-RAY DIFFRACTIONf_dihedral_angle_d14.241565
X-RAY DIFFRACTIONf_chiral_restr0.038233
X-RAY DIFFRACTIONf_plane_restr0.004253
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
1.9521-2.00490.26621280.24771167X-RAY DIFFRACTION91
2.0049-2.06390.26591410.21861295X-RAY DIFFRACTION98
2.0639-2.13050.26271360.2061263X-RAY DIFFRACTION98
2.1305-2.20670.24241500.20261287X-RAY DIFFRACTION98
2.2067-2.2950.23141380.20061297X-RAY DIFFRACTION99
2.295-2.39950.2591450.20011290X-RAY DIFFRACTION99
2.3995-2.5260.23581450.19761288X-RAY DIFFRACTION100
2.526-2.68420.30041440.19921309X-RAY DIFFRACTION100
2.6842-2.89140.27621420.20831317X-RAY DIFFRACTION100
2.8914-3.18240.28991500.22121329X-RAY DIFFRACTION100
3.1824-3.64270.24431440.20051317X-RAY DIFFRACTION99
3.6427-4.58890.19251480.16311327X-RAY DIFFRACTION98
4.5889-47.980.21151580.18111410X-RAY DIFFRACTION100
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)
11.76510.01580.52354.9425-1.98054.53160.1949-0.65330.14260.6243-0.0920.5776-0.2869-0.5104-0.08390.4574-0.00880.10420.4285-0.04670.3159-13.7942-13.402313.6926
23.89921.3044-1.04173.1032-1.06554.2259-0.5065-0.52410.0103-0.1002-0.0825-0.84650.30990.01470.56740.39150.0585-0.04210.49310.0410.5002-4.665-16.574511.618
38.8904-3.39643.1122.8402-0.54725.1035-0.0718-1.08820.6009-0.6085-0.1369-1.3209-0.629-0.5102-0.17730.9637-0.2122-0.0110.62130.11580.54931.493-3.48232.0746
49.5612-1.44881.70012.02174.08183.9692-0.21720.8941-1.11470.3409-0.0976-1.33520.14931.0870.12870.2989-0.0022-0.00660.5798-0.05560.3433-0.3662-15.98770.0144
53.35311.10840.94454.15920.91946.5359-0.2398-0.2146-0.29140.05120.31990.33830.2668-0.4292-0.0090.27960.03670.06560.31850.02140.3481-13.3927-18.28894.137
64.7331-5.9794-0.66387.41030.81370.5735-0.06310.6602-0.1614-0.1419-0.51690.90.5891-0.7312-0.51450.5311-0.09120.03780.3803-0.10980.6333-20.14997.00340.0664
72.89420.4103-0.32042.6344-0.23190.7677-0.12670.30620.117-0.25640.06010.163-0.0968-0.09060.09280.3779-0.0292-0.00130.3896-0.04460.3659-13.7197-20.0403-15.994
82.13430.31131.20130.6411-0.66812.00140.2309-0.92460.27430.86580.16120.3366-0.6564-0.00170.15320.3885-0.0593-0.03760.44590.01180.7218-20.1167-32.6585-4.8256
94.6272-1.66222.11937.903-0.42144.8356-0.09930.1345-0.1729-0.02230.11140.15-0.27570.1048-0.06610.285-0.02670.00150.2696-0.06380.2817-10.146-16.4939-7.5662
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection details
1X-RAY DIFFRACTION1chain 'A' and (resid 1 through 20 )
2X-RAY DIFFRACTION2chain 'A' and (resid 21 through 42 )
3X-RAY DIFFRACTION3chain 'A' and (resid 43 through 48 )
4X-RAY DIFFRACTION4chain 'A' and (resid 49 through 55 )
5X-RAY DIFFRACTION5chain 'A' and (resid 56 through 88 )
6X-RAY DIFFRACTION6chain 'A' and (resid 89 through 96 )
7X-RAY DIFFRACTION7chain 'B' and (resid 1 through 42 )
8X-RAY DIFFRACTION8chain 'B' and (resid 43 through 50 )
9X-RAY DIFFRACTION9chain 'B' and (resid 51 through 86 )

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