Poster abstracts

Poster number 77 submitted by Lexie Kuzmishin

Function of a putative trans-editing factor, ProXp-x, in Rhodopseudomonas palustris

Lexie Kuzmishin (The Ohio State Biochemistry Program, Department of Chemistry and Biochemistry, and Center for RNA Biology, The Ohio State University), Ziwei Liu (Department of Chemistry and Biochemistry, and Center for RNA Biology, The Ohio State University), Jo Marie Bacusmo (Department of Chemistry and Biochemistry, and Center for RNA Biology, The Ohio State University), Karin Musier-Forsyth (Department of Chemistry and Biochemistry, and Center for RNA Biology, The Ohio State University)

Abstract:
Aminoacyl-tRNA synthetases (ARSs) attach specific amino acids to cognate tRNAs; this process is a critical checkpoint in protein synthesis. Due to the similarity in size and structure of many amino acids, ARSs are known to misactivate both proteinogenic and non-protein amino acids. Thus, editing mechanisms are needed to prevent widespread mistranslation. In addition to specific tRNAPro editing mediated by the insertion (INS) domain present in most bacterial prolyl-tRNA synthetases (ProRS), single-domain trans-editing factors structurally homologous to INS have evolved in some organisms. To date, INS-like trans-editing proteins have been shown to act on specific tRNAs mischarged with proteogenic amino acids. Here, we show that Rhodopseudomonas palustris ProXp-x, a previously uncharacterized INS homolog, displays robust editing of tRNAs mischarged with the non-protein amino acids α-aminobutyrate (2-Abu) and D-Thr in vitro. In vivo experiments have demonstrated that 2-Abu is toxic to an E. coli strain encoding an editing-defective valyl-tRNA synthetase, and expression of R. palustris ProXp-x rescues cell growth. Similarly, preliminary in vivo experiments indicate D-Thr is toxic to E. coli when overexpressing threonyl-tRNA synthetase, and co-expression of R. palustris ProXp-x can rescue the growth defect . An R. palustris ProXp-x deletion strain has been constructed and experiments are underway to characterize growth phenotypes under various conditions . Metabolomics studies are also in progress to further understand which environmental conditions cause intracellular accumulation of 2-Abu and D-Thr . Based on our results, we hypothesize that editing by ProXp-x may mitigate misincorporation of non-protein amino acids under environmental conditions or stresses in which non-protein amino acids pose a greater threat to the cell.

Keywords: trans-editing, aminoacyl-tRNA synthetases, Rhodopseudomonas palustris