Abstract
Prokaryotic Toxin-Antitoxin (TA) systems typically consist of a toxin and antitoxin pair encoded by a small operon. When activated, a TA system will transition from a neutralised to an active state, in which the antitoxin ceases to inhibit the toxin. Active toxin will typically modify a conserved cellular protein and halt an essential cellular process, causing cell death or dormancy. While the function of TA systems is frequently unclear, they have increasingly demonstrated immune defence against the viruses of bacteria, bacteriophages (phages). Previously, we found that the operon mmpAB formed a functional TA system. Here, we sought to improve our understanding of mmpAB, focusing on the mechanism of MmpB toxicity, how MmpA provided toxin-neutralisation, the trigger of activation, and what function mmpAB provided to the host cell. Through structural predictions, functional assays, microscopy, and flow cytometry, we found that MmpB likely catalysed the addition of AMP to a conserved cellular target, similar to but distinct from other Fic-domain toxins. Through structural predictions and functional assays, we found that MmpA neutralised MmpB analogously to canonical Fic-associated antitoxins but with a novel motif. Further, we found evidence supporting MmpA-mediated transcriptional autoregulation, previously absent in Fic-based toxin-antitoxin systems. Crucially, mmpAB defended against Tequintavirus Φ34, in a manner dependent on the Fic domain of MmpB. Strains of Φ34 that evaded mmpAB defence contained mutations in the essential pre-early protein A1, implicating this protein in mmpAB defence, and furthering our understanding of the two-step infection process unique to Tequintavirus species. In summary, these findings establish a connection between Fic toxins and canonical TA biology, and further demonstrate the role that TA systems play in phage defence. Consequently, mmpAB may play a future role in the inhibition or enhancement of bacteriophage in managing bacterial cell populations.