Poster abstracts
Poster number 24 submitted by Lexie Kuzmishin
Quality control by trans-editing factor prevents global mistranslation of non-protein amino acids
Alexandra B. Kuzmishin (OSBP, Department of Chemistry and Biochemistry, Center for RNA Biology, The Ohio State University), Jo Marie Bacusmo, William A. Cantara (Department of Chemistry and Biochemistry, Center for RNA Biology, The Ohio State University), Yuki Goto, Hiroaki Suga (Department of Chemistry, Graduate School of Science, The University of Tokyo), Karin Musier-Forsyth (Department of Chemistry and Biochemistry, Center for RNA Biology, The Ohio State University)
Abstract:
Accuracy in protein synthesis is maintained through multiple pathways, with a critical checkpoint occurring at the tRNA aminoacylation step catalyzed by aminoacyl-tRNA synthetases (AARSs). In addition to Ala-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 are present in some organisms. To date, INS-like editing proteins have been shown to act on specific tRNAs mischarged with genetically encoded amino acids. However, structurally related non-protein amino acids are ubiquitous in cells and threaten the proteome. Here, we show that Rhodopseudomonas palustris ProXp-x, a previously uncharacterized INS homolog, edits a known ProRS aminoacylation error, Ala-tRNAPro, but displays even more robust editing of tRNAs misaminoacylated with the non-protein amino acid α-aminobutyrate (Abu) in vitro and in vivo. Additionally, ProXp-x robustly edits D-Ala-tRNAPro and D-Thr-tRNAVal, but shows no activity towards L-Thr in vitro. Abu is the product of the transamination of oxobutyrate and is a precursor metabolite in Thr biosynthesis, but D-amino acids are synthesized from their L-isomers or are acquired from the environment or surrounding organisms. Abu is mischarged by E. coli ValRS and ProRS in vitro, and in vivo experiments showed that Abu is toxic to an E. coli strain encoding an editing-defective ValRS (1). However, expression of R. palustris ProXp-x rescues cell growth, presumably by removing Abu that has been mischarged onto tRNAVal by the editing-deficient ValRS. Our results indicate that editing by ProXp-x may offer advantages to cells, especially under environmental conditions where concentrations of non-protein amino acids may challenge the substrate specificity of AARSs. Cell-based experiments are underway to explore phenotypic differences between wild-type R. palustris and a ProXp-x null R. palustris strain under a variety of growth conditions.
References:
1. Döring V, Mootz HD, Nangle LA, Hendrickson TL, de Crécy-Lagard V, Schimmel P, Marlière P. Enlarging the amino acid set of Escherichia coli by infiltration of the valine coding pathway. Science. 2001 April 20; 292:501-4.
Keywords: trans-editing, ProXp-x, INS