Viral Genome Delivery

The goal of this research is to understand how double stranded DNA viruses deliver their large (50-250kbs) genomes into infected cells. In tailed bacteriophages and Herpesviruses, this process is mediated by a large 12-fold symmetric oligomeric protein, named portal protein. The portal protein forms the conduit throughout which viral DNA is ejected from the capsid into the host and is thought to function as a dedicated DNA pump. Most of the work carried out in my laboratory is based on a simple viral model, the bacteriophage P22.

P22 is a tailed phage of the Podoviridae family that infects Salmonella enterica. The delivery of P22 genetic information (~50kbs) through the Salmonella cell envelope is extremely efficient and occurs in 60-90 seconds. Such a remarkable example of biotransformation is mediated by the virus tail apparatus, a 2.8MDa molecular machine built by five proteins repeated in various stoichiometries (Figure 1). In collaboration with Dr. Sherwood Casjens, at the University of Utah, we have isolated and reconstituted the five-polypeptide chains forming the phage P22 tail. Using a wide repertoire of structural (e.g. X-ray crystallography, electron cryo-microscopy), biochemical and virological techniques, we have determined the structure, composition, and assembly of this macromolecular machine.

Figure 1. Phage P22 DNA injecting machine. The DNA injection apparatus is in green (from Lander et al, Science 2007). The portal protein gp1 provides a surface of attachment for the tail apparatus formed by gp9, gp4, gp10, and gp26 (magnified on the right panel).


We recently determined the crystal structure of P22 tail needle gp26 at 2.1Å resolution (pdb 2POH, Olia et al., Nature Struc Mol Biol. 2007). Gp26 consists is homotrimer protein fiber ~240Å in length and only ~25Å in diameter, which has striking resemblance to a drill bit . Gp26 displays remarkable structural stability.

Figure 2. Ribbon diagram of Tail Needle gp26 (Olia et al., Nature Struc Mol Biol. 2007).


Our current work is aimed at defining how P22 delivers its genome into the host. More specifically, we are investigating three steps of the DNA delivery reaction:

  1. the penetration of the bacterial cell envelope, which is mediated by the tail needle gp26,
  2. the mechanisms of DNA ejection through the host cell envelope, which is mediated by the portal protein,
  3. the role of P22 injection proteins gp7, gp16 and gp20, which are required for infectivity.

We are also interested in visualizing the structure of the 12-fold symmetric portal proteins both in bacteriophages and Herpesviruses. In collaboration with Dr. Tamir Gonen, at the University of Washington, we recently determined the medium resolution structure of P22 portal protein (Fig .3).

For phage P22 portal protein, we recently obtained crystal of the truncated P22 portal protein in complex with 12 copies of tail factor gp4. Despite the large size of this macromolecular complex (M.W. 1.1MDa), and the fact that the asymmetric unit contains two portal:gp4 complexes, complete diffraction data to ~3.3Å resolution have been accurately collected. A characteristic diffraction pattern is shown below.