Poster abstracts
Poster number 12 submitted by Lexie Kuzmishin Nagy
Towards the Characterization of a Bacterial D-Aminoacyl-tRNA Trans-Editing Enzyme
Lexie Kuzmishin Nagy (Department of Biology, Department of Chemistry, Earlham College), Dawt Sang (Department of Biology, Department of Chemistry, Earlham College), Ngoc Tuong Vy Nguyen (Department of Biology, Department of Chemistry, Earlham College), Landry Cox (Department of Biology, Department of Chemistry, Earlham College), Samia Kassim (Department of Biology, Department of Chemistry, Earlham College)
Abstract:
Correctly synthesized aminoacyl-tRNAs (aa-tRNAs) are imperative for accurate protein synthesis. Mischarged aminoacyl-tRNAs can deliver incorrect amino acids to the ribosome during translation, leading to amino acid misincorporation in the nascent. Aminoacyl-tRNA synthetases (aaRSs) can mischarge tRNAs with non-cognate L-amino, however some aaRSs also mischarge D-amino acids onto tRNAs. D-amino acids, which are highly abundant in soil, stall ribosomes and few editing enzymes remove mischarged D-amino acids from tRNAs. As a result, D-aa-tRNAs accumulate in cells, depleting the pool of free tRNA available for use in protein synthesis. Most organisms encode D-aa-tRNA deacylase enzymes (DTDs) that hydrolyze D-aa-tRNAs to prevent their accumulation and the resulting D-amino acid toxicity. However, soil-dwelling rhizobia bacteria do not encode DTDs, but do encode ProXp-x, a single-domain enzyme homologous to the Prolyl-tRNA synthetase aa-tRNA editing domain. Rhodopseudomonas palustris ProXp-x was previously shown to edit Ala-tRNAPro and tRNAs mischarged with the non-protein amino acid &alpha-aminobutyrate. Preliminary work demonstrates ProXp-x also hydrolyzes certain combinations of D-aminoacyl-tRNAs in vitro with a substrate selection mechanism distinct from DTD. ProXp-x recognizes both amino acid enantiomers, unlike DTD, and edits various, unrelated mischarged tRNA isoacceptors. Previous work suggests that both the amino acid moiety and tRNA moiety contribute to substrate recognition. We used deacylation assays and binding assays to study ProXp-x’s selection of tRNA and amino acid enantiomers. Preliminary data suggest ProXp-x has low affinity for deacylated, wildtype tRNAs. We also identified a viability-related D-amino acid toxicity phenotype in an Escherichia coli Δdtd strain. Taken together, their preliminary data suggest ProXp-x is a putative D-aminoacyl-tRNA deacylase in rhizobia that may help these bacteria respond to the threat of D-amino acids in their environment.
Keywords: tRNA, non-protein amino acids, editing