Integrative Structural Analysis of Phage DEV Reveals a Genome Ejection Motor

DEV is an obligatory lytic phage of the N4-like genus, recently reclassified as . The DEV genome encodes 91 ORFs, including a 3,398 amino acid virion-associated RNA polymerase. Here, we describe the complete architecture of DEV, determined using a combination of cryo-electron microscopy localized reconstruction, biochemical methods, and genetic knockouts. We built de structures of all capsid factors and tail components involved in host attachment. We demonstrate that DEV long tail fibers are essential for infection of and dispensable for infecting mutants with a truncated lipopolysaccharide devoid of the O-antigen. We identified DEV ejection proteins and, unexpectedly, found that the giant DEV RNA polymerase, the hallmark of the family, is an ejection protein. We propose that DEV ejection proteins form a genome ejection motor across the host cell envelope and that these structural principles are conserved in all .