Poster abstracts
Poster number 63 submitted by Lexie Kuzmishin
Function of a putative trans-editing factor, ProXp-x, in Rhodopseudomonas palustris
Lexie Kuzmishin (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, these enzymes are known to misactivate both proteinogenic and non-protein amino acids. Thus, editing mechanisms are needed to prevent widespread mistranslation. In addition to editing mediated by inserted (INS) domains present in some 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 genetically encoded amino acids. However, structurally related non-protein amino acids also threaten the proteome. 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, 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. Overall, our results suggest that editing by trans-editing domains such as ProXp-x may mitigate misincorporation of non-protein amino acids under environmental conditions or stresses in which fluctuating metabolic pools pose a greater threat to the cell.
Keywords: aminoacyl-tRNA, editing, synthetase