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φ 29

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

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φ 29

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  1. Tailed bacteriophages: Structure, Function and Evolution(A tale of two tails)Michael RossmannPurdue University

  2. φ29 T4

  3. 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 ~20 ORFs 8 structural proteins 1 structural RNA Host: B. Subtilis Gram positive Cell wall external to single cell membrane Packeges DNAgp3 genome

  4. Bacteriophage phi29 Rossmann Lab Valorie Bowman Paul Chipman Kay Choi Marc Morais Alan Simpson Yizhi Tao Ye Xiang Anderson Lab Mohammed Badasso Shelley Grimes Paul Jardine Jaya Kotl

  5. Tao et al Cell, 1998, 95:431-437

  6. Three layers of DNA DNA ring Connector appendages gp3? head fiber bulge lower collar knob ?

  7. Phi29 head-tail head-tail Connector

  8. φ29 dodecameric connector Simpson et al,Nature, 2000,408:745-750

  9. pRNA difference map

  10. Partially packaged prohead with pRNA and ATPase (gp16) Empty Prohead with pRNA

  11. Connector pRNA pentamer Difference density Gp16 ATPase

  12. Phage Φ29: DNA packaging motor Simpson et al,Nature, 2000,408:745-750

  13. The T4 structure Purdue University Anthoni Batisti Paul Chipman Andrei Fokine Victor Kostyuchenko Pertr Leiman Moscow Vadim Mesyanzhinov Mikhail Schneider Catholic University of America Venigella Rao Tokyo Institute of Technology Fumio Arisaka Shuji Kanamaru NIH Bethesda Bijan Ahvazi Karen Boeshans Alasdair Steven University of Maryland Lindsay Black Santiago Mark van Raaij

  14. Too complex to crystallize: Bacteriophage T4an efficient infection machine

  15. Phage particle assembly Head Long tail fibers Fibritin gp13, gp14, [gp2], [gp4] [gp63] Tail

  16. Tail terminator gp15 138 gp18 138 gp19 T4 Tail assembly Wedge 6 25, 6 53, 7, 8 10, 11 gp9 15S gp12 gp48 gp54 120S Hub 70S 5, 27 29 22S

  17. Crystal structures of the baseplate proteins gp9, LTF attachment gp11, STF attachment gp8 gp12 C-term gp12, STF gp5-gp27 30 nm

  18. Common topologies gp11 gp10 gp12 Interaction in 3D X big insertion

  19. 80 Å Gp5-gp27 complex structure overview gp27 trimer gp5 N-terminal domain 190 Å gp5 lysozymedomain gp5 C-terminal domain Kanamaru et al, Nature 2002,415:553-557

  20. 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 Å resolution

  21. Baseplate movie 19 48 54 6 25 53 9 27 8 7 5 10 11 12 26? Kostyuchenko et al, Nat. Struct. Biol. 2003,10:688-693

  22. Hexagonal Baseplate

  23. Star-shaped baseplates (contracted tails) • Sixfold symmetry • 1580 particles used in the reconstruction • Defoci 0.7 – 3.3 mm • 17 Å resolution Leiman et al, Cell 2004, 118:419-429

  24. Contracted tail Structure Contracted tail movie 23 20 13 14 15 3 19 18 34 9 7 8 6 25 53 11 10 12 19

  25. Extended sheath 1 turn Contracted sheath 2 turns Kostyuchenko et al, Nat.Struct, Mol. Biol. 2005 in press

  26. November 18th, 2004

  27. Movie

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