FhuA-mediated phage genome transfer into liposomes: A cryo-electron tomography study

Background: The transfer of phage genomes into host cells is a well establi shed but only dimly understood process. Following the irreversible phage bi nding to a receptor in the bacterial outer membrane, the DNA is ejected fro m the viral capsid and transferred across the bacterial cell envelope. In E scherichia coli, the mere interaction of the phage T5 with its outer membra ne receptor, the ferrichrome transporter FhuA, is sufficient to trigger the release of the DNA from the phage capsid. Although the structure of FhuA h as been determined at atomic resolution, the understanding of the respectiv e roles of phage and bacterial proteins in DNA channeling and the mechanism s by which the transfer of the DNA is mediated remains fragmentary. Results: We report on the use of cryo-electron tomography to analyze, at a molecular level, the interactions of T5 phages bound to FhuA-containing pro teoliposomes. The resolution of the three-dimensional reconstructions allow ed us to visualize the phage-proteoliposome interaction before and after re lease of the genome into the vesicles. After binding to its receptor, the s traight fiber of the phage T5 (the "tip" of the viral tail made of pb2 prot eins) traverses the lipid bilayer, allowing the transfer of its double-stra nded DNA (121,000 bp) into the proteoliposome. Concomitantly, the tip of th e tail undergoes a major conformational change; it shrinks in length (from 50 to 23 nm), while its diameter increases (from 2 to 4 nm). Conclusions: Taking into account the crystal structure of FhuA, we conclude that FhuA is only used as a docking site for the phage. The tip of the pha ge tail acts like an "injection needle," creating a passageway at the perip hery of FhuA, through which the DNA crosses the membrane. A possible mechan istic scenario for the transfer of the viral genome into bacteria is discus sed.