Structure of epsilon15 bacteriophage reveals genome organization and DNA packaging/injection apparatus

Abstract

The critical viral components for packaging DNA, recognizing and binding to host cells, and injecting the condensed DNA into the host are organized at a single vertex of many icosahedral viruses. These component structures do not share icosahedral symmetry and cannot be resolved using a conventional icosahedral averaging method. Here we report the structure of the entire infectious Salmonella bacteriophage epsilon15 (ref. 1) determined from single-particle cryo-electron microscopy, without icosahedral averaging. This structure displays not only the icosahedral shell of 60 hexamers and 11 pentamers, but also the non-icosahedral components at one pentameric vertex. The densities at this vertex can be identified as the 12-subunit portal complex sandwiched between an internal cylindrical core and an external tail hub connecting to six projecting trimeric tailspikes. The viral genome is packed as coaxial coils in at least three outer layers with 90 terminal nucleotides extending through the protein core and the portal complex and poised for injection. The shell protein from icosahedral reconstruction at higher resolution exhibits a similar fold to that of other double-stranded DNA viruses including herpesvirus2,3,4,5,6, suggesting a common ancestor among these diverse viruses. The image reconstruction approach should be applicable to studying other biological nanomachines with components of mixed symmetries.

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Figure 1: Structure of epsilon15 bacteriophage.
Figure 2: The tail structure.
Figure 3: Structure of the portal complex and internal core.
Figure 4: Shell protein structure.

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Acknowledgements

We acknowledge the support of grants from National Institutes of Health and the Robert Welch Foundation. We thank M. Dougherty for the production of the animations, M. Baker for the AIRS program for secondary structure element identification, and M. F. Schmid and F. Rixon for discussions. Author Contributions W.J. developed the image processing methods and solved and analysed the structures; J.C. and J.J. collected the 200- and 300-kV image data respectively; P.W. did the biochemical preparation and analysis; and W.J., P.W., J.K. and W.C. interpreted the structure and wrote the manuscript.

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Correspondence to Wah Chiu.

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Competing interests

The three-dimensional density maps have been deposited into the EBI-MSD EMD database with accession codes EMD-1175 for the complete structure without symmetry imposition and EMD-1176 for the icosahedral shell structure. Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Supplementary information

Supplementary Methods

This file contains additional details on the methods used in this study, including purification of Epsilon15 phage, CryoEM imaging, 3-D icosahedral reconstruction 3-D non-icosahedral reconstruction and a structural analysis. (DOC 46 kb)

Supplementary Figures

Supplementary Figures 1–8 with accompanying legends. (PPT 12200 kb)

Supplementary Movie 1

The complete structure of Epsilon15 phage showing each of the structural components including tailspikes, tail hub, portal, core, dsDNA and shell proteins based on the 20 Å reconstruction with no symmetry imposed. (MPG 33850 kb)

Supplementary-Movie 2

The icosahedral reconstruction of Epsilon15 phage at 9.5 Å resolution showing the locations of the α-helices and β-sheets of the average capsid shell protein. (MPG 35257 kb)

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