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Tailed bacteriophages: Structure, Function and Evolution (A tale of two tails) Michael Rossmann Purdue University. φ 29. T4. T4 168 kbp genome ~300 ORFs 40 structural proteins Host: E. Coli Gram negative Cell wall inside periplasmic space Packages T4 concatomer. Phi29 20 kbp genome - PowerPoint PPT Presentation

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  • Tailed bacteriophages: Structure, Function and Evolution

    (A tale of two tails)

    Michael RossmannPurdue University

  • T429

  • T4168 kbp genome

    ~300 ORFs

    40 structural proteins

    Host: E. Coli

    Gram negativeCell wall inside periplasmic space

    Packages T4 concatomerPhi2920 kbp genome

    ~20 ORFs

    8 structural proteins1 structural RNA

    Host: B. Subtilis

    Gram positiveCell wall external to single cell membrane

    Packeges DNAgp3 genome

  • Bacteriophage phi29Rossmann LabValorie BowmanPaul ChipmanKay ChoiMarc MoraisAlan SimpsonYizhi TaoYe XiangAnderson LabMohammed BadassoShelley GrimesPaul JardineJaya Kotl

  • Tao et al Cell, 1998, 95:431-437

  • gp3?head fiberbulge lower collarknobDNA ring

    Connector

    appendages?Three layers of DNA

  • Phi29 head-tail head-tail Connector

  • Simpson et al, Nature, 2000, 408:745-750 29 dodecameric connector

  • pRNA difference map

  • Partially packaged proheadwith pRNA and ATPase (gp16)Empty Proheadwith pRNA

  • ConnectorpRNA pentamerDifference densityGp16 ATPase

  • Phage 29:DNA packaging motorSimpson et al, Nature, 2000, 408:745-750

  • The T4 structure

    Purdue UniversityAnthoni BatistiPaul ChipmanAndrei FokineVictor KostyuchenkoPertr LeimanMoscowVadim MesyanzhinovMikhail SchneiderCatholic University of AmericaVenigella RaoTokyo Institute of TechnologyFumio ArisakaShuji KanamaruUniversity of MarylandLindsay BlackNIH BethesdaBijan AhvaziKaren BoeshansAlasdair StevenSantiago Mark van Raaij

  • Too complex to crystallize: Bacteriophage T4 an efficient infection machine

  • Phage particle assemblyHeadTailFibritingp13, gp14, [gp2], [gp4][gp63]Long tail fibers

  • T4 Tail assembly5, 2753, 7, 810, 1125, 615S2922SWedgeHub70S6120Sgp9gp12gp48gp54

  • Crystal structures of the baseplate proteinsgp8gp11,STF attachmentgp9,LTF attachmentgp5-gp27gp12C-term30 nmgp12, STF

  • Common topologiesgp11gp10gp12Interaction in 3DXbig insertion

  • Gp5-gp27 complex structure overviewgp27 trimergp5 N-terminal domaingp5 lysozyme domaingp5 C-terminaldomain80 190 Kanamaru et al, Nature 2002, 415:553-557

  • Hexagonal conformation (tube-baseplates)Initial model hexagonal prism connected to a tube

    Sixfold symmetry

    945 particles used in the reconstruction

    Defoci 1.5 3.5 mm

    12 resolution

  • Baseplate movie1948 546 25 53987101112526?27Kostyuchenko et al, Nat. Struct. Biol. 2003, 10:688-693

  • Hexagonal Baseplate

  • Star-shaped baseplates (contracted tails)Sixfold symmetry

    1580 particles used in the reconstruction

    Defoci 0.7 3.3 mm

    17 resolution

    Leiman et al, Cell 2004, 118:419-429

  • Contracted tail movie19189871011122334152013 143196 25 53Contracted tailStructure

  • Extended sheath1 turnContracted sheath2 turnsKostyuchenko et al, Nat.Struct, Mol. Biol. 2005 in press

  • November 18th, 2004

  • Movie

    T4 + phi29Phi29 assemblyDiagram of phi29 with dimensionspRNA difference mapPartially filled prohead with gp16 ATPase