Structural dynamics of bacteriophage P22 infection initiation revealed by cryo-electron tomography

Abstract

SUMMARY For successful infection, bacteriophages must overcome multiple barriers to transport the genome and proteins across the bacterial cell envelope. We use cryo-electron tomography to study infection initiation of phage P22 in Salmonella enterica sv. Typhimurium, revealing how a channel forms to allow genome translocation into the cytoplasm. Our results show free phages initially attaching obliquely to the cell through interactions between the O antigen and two of the six tailspikes; the tail needle also abuts the cell surface. The virion then orients to the perpendicular and the needle penetrates the outer membrane. The needle is released and the internal head protein gp7* is ejected and assembles into an extra-cellular channel extending from the gp10 baseplate to the cell surface. A second protein, gp20, is ejected and assembles into a structure that extends the extra-cellular channel across the outer membrane into the periplasm. Insertion of the third ejected protein gp16 into the cytoplasmic membrane likely completes the overall trans-envelope channel into the cytoplasm. Construction of a trans-envelope channel is an essential step during infection by all short-tailed phages of Gram-negative bacteria because such virions cannot directly deliver their genome into the cell cytoplasm.

Phage diversity, genomics and phylogeny

“…Members of the Podoviridae family, such as E. coli phage T7, have very short and non-contractile tails. A tube-like extension of the tail that penetrates both cell membranes was observed to be essential for genome delivery into the host 29 .…”

Increasing the Affinity of an O‐Antigen Polysaccharide Binding Site in Shigella flexneri Bacteriophage Sf6 Tailspike Protein

“…Binding the multivalent TSP to the multivalent sensor surface mimics the situation during phage infection, where the TSP as part of the phage tail has to bind perpendicular to the bacterial cell surface. Although the also perpendicularly protruding O‐antigen chains would suggest a mainly parallel orientation of TSP and polysaccharide chains, cryoelectron tomography analyses showed that whole phage particles bound obliquely to the cell surface . Therefore, an SPR surface‐immobilized polysaccharide that might contain also surface‐parallel regions mimics this situation, as periodate treatment of the polysaccharide might interfere with its fully perpendicular attachment.…”

Increasing the Affinity of an O-Antigen Polysaccharide Binding Site in Shigella Flexneri Bacteriophage Sf6 Tailspike Protein

“…Althought he also perpendicularly protruding O-antigen chains would suggest a mainly parallel orientation of TSP and polysaccharide chains, cryoelectron tomography analysess howedt hat whole phage particles bound obliquelyt ot he cell surface. [70] Therefore, an SPR surface-immobilized polysaccharide that might contain also surface-parallel regions mimics this situation, as periodate treatment of the polysaccharide might interfere with its fully perpendicular attachment. Here, interaction with more than one glycan binding site per TSP would then require a6 0 8 rotation aroundt he symmetry axis and result in an energetic penalty.I ndeed, the negative cooperativity found in the Scatchard plot might point to this unfavourable surface binding where two sites on one TSP must bind simultaneously.M odelst hat are more sophisticated should be therefore used that take into account the reactionv olumea nd ap robability factor for the multivalent bindinge vent.…”