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Bacteriophage 80α is a representative of a class of temperate phages that infect Staphylococcus aureus and other Gram-positive bacteria. Many of these phages carry genes encoding toxins and other virulence factors. This phage, 80α, is also involved in high-frequency mobilization of S. aureus pathogenicity islands (SaPIs), mobile genetic elements that carry virulence factor genes. Bacteriophage 80α encodes a minor capsid protein, gp44, between the genes for the portal protein and major capsid protein. Gp44 is essential for a productive infection by 80α but not for transduction of SaPIs or plasmids. We previously demonstrated that gp44 is an ejection protein that acts to promote progression to the lytic cycle upon infection and suggested that the protein might act as an anti-repressor of CI in the lytic-lysogenic switch. However, an 80α Δ44 mutant also exhibited a reduced rate of lysogeny. Here, we show that gp44 is a non-specific DNA binding protein with affinity for the blunt ends of linear DNA. Our data suggest a model in which gp44 promotes circularization of the genome after injection into the host cell, a key initial step both for lytic growth and for the establishment of lysogeny.

In the tailed bacteriophages, DNA is packaged into spherical procapsids, leading to expansion into angular, thin-walled mature capsids. In many cases, this maturation is accompanied by cleavage of the major capsid protein (CP) and other capsid-associated proteins, including the scaffolding protein (SP) that serves as a chaperone for the assembly process. Staphylococcus aureus bacteriophage 80α is capable of high frequency mobilization of mobile genetic elements called S. aureus pathogenicity islands (SaPIs), such as SaPI1. SaPI1 redirects the assembly pathway of 80α to form capsids that are smaller than those normally made by the phage alone. Both CP and SP of 80α are N-terminally processed by a host-encoded protease, Prp. We have analyzed phage mutants that express pre-cleaved or uncleavable versions of CP or SP, and show that the N-terminal sequence in SP is absolutely required for assembly, but does not need to be cleaved in order to produce viable capsids. Mutants with pre-cleaved or uncleavable CP display normal viability. We have used cryo-EM to solve the structures of mature capsids from an 80α mutant expressing uncleavable CP, and from wildtype SaPI1. Comparisons with structures of 80α and SaPI1 procapsids show that capsid maturation involves major conformational changes in CP, consistent with a release of the CP N-arm by SP. The hexamers reorganize during maturation to accommodate the different environments in the 80α and SaPI1 capsids., Competing Interests: The authors declare no conflict of interest.