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Cell Rep. 2014 Apr 24;7(2):339-47. doi: 10.1016/j.celrep.2014.03.034. Epub 2014 Apr 13.

Structure, mechanism, and specificity of a eukaryal tRNA restriction enzyme involved in self-nonself discrimination.

Author information

1
Molecular Biology Program, Sloan-Kettering Institute, New York, NY 10065, USA.
2
Molecular Biology Program, Sloan-Kettering Institute, New York, NY 10065, USA. Electronic address: s-shuman@ski.mskcc.org.

Abstract

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 tRNA(Gln(UUG)). We find that recombinant PaT incises a synthetic tRNA(Gln(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.

PMID:
24726365
PMCID:
PMC4121121
DOI:
10.1016/j.celrep.2014.03.034
[Indexed for MEDLINE]
Free PMC Article

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