| Summary: | Summary: tRNA restriction by anticodon nucleases underlies cellular stress responses and self-nonself discrimination in a wide range of taxa. Anticodon breakage inhibits protein synthesis, which, in turn, results in growth arrest or cell death. The eukaryal ribotoxin PaT secreted by Pichia acaciae inhibits growth of Saccharomyces cerevisiae via cleavage of tRNAGln(UUG). We find that recombinant PaT incises a synthetic tRNAGln(UUG) stem-loop RNA by transesterification at a single site 3′ of the wobble uridine, yielding 2′,3′-cyclic phosphate and 5′-OH ends. Incision is suppressed by replacement of the wobble nucleobase with adenine or guanine. The crystal structure of PaT reveals a distinctive fold and active site, essential components of which are demonstrated by mutagenesis. Pichia acaciae evades self-toxicity via a distinctive intracellular immunity protein, ImmPaT, which binds PaT and blocks nuclease activity. Our results highlight the evolutionary diversity of tRNA restriction and immunity systems. : tRNA restriction via incision of the anticodon loop is a deeply rooted response to cellular stress and plays a prominent role in self-nonself discrimination by microbes. Here, Shuman and colleagues report the crystal structure of Pichia acaciae toxin PaT, a eukaryal tRNA restriction enzyme that breaks the tRNAGln(UUG) anticodon stem-loop 3′ of the wobble uridine. PaT has a unique fold and active site. Pichia acaciae evades self-toxicity via an immunity protein, ImmPaT, that binds PaT and effaces nuclease activity.
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