A viral protein antibiotic inhibits lipid II flippase activity

For bacteriophage infections, the cell walls of bacteria, consisting of a single highly polymeric molecule of peptidoglycan (PG), pose a major problem for the release of progeny virions. Phage lysis proteins that overcome this barrier can point the way to new antibacterial strategies(1), especially small lytic single-stranded DNA (the microviruses) and RNA phages (the leviviruses) that effect host lysis using a single non-enzymatic protein(2). Previously, the A(2) protein of levivirus Q beta and the E protein of the microvirus phi X174 were shown to be 'protein antibiotics' that inhibit the MurA and MraY steps of the PG synthesis pathway(2-4). Here, we investigated the mechanism of action of an unrelated lysis protein, Lys(M), of the Escherichia coli levivirus M-5. We show that Lys(M) inhibits the translocation of the final lipid-linked PG precursor called lipid II across the cytoplasmic membrane by interfering with the activity of MurJ. The finding that Lys(M) inhibits a distinct step in the PG synthesis pathway from the A(2) and E proteins indicates that small phages, particularly the single-stranded RNA (ssRNA) leviviruses, have a previously unappreciated capacity for evolving novel inhibitors of PG biogenesis despite their limited coding potential.