Poster abstracts
Poster number 22 submitted by Jun-Kyu Byun
Role of Unique C-terminal Domain in an Aminoacyl-tRNA Trans-editing Protein from Arabidopsis thaliana
Jun-Kyu Byun (Department of Chemistry and Biochemistry, Center for RNA Biology, Ohio State University), Marina Bakhtina (Department of Chemistry and Biochemistry, Center for RNA Biology, Ohio State University), Karin Musier-Forsyth (Department of Chemistry and Biochemistry, Center for RNA Biology, Ohio State University)
Abstract:
Aminoacyl-tRNA synthetases (ARSs) play a central role in maintaining high fidelity of protein synthesis, which is essential for cell viability. Due to the structural similarity of amino acids, some ARSs are error prone and mispair non-cognate amino acids with specific tRNA. Thus, to prevent mistranslation, quality control mechanisms are required. Prolyl-tRNA synthetase (ProRS) can misactivate and mischarge noncognate alanine to form Ala-tRNA-Pro. Some bacterial ProRSs possess an editing domain that can deacylate Ala-tRNA-Pro, but some bacteria and all eukaryotes lack this editing domain in the context of ProRS. Instead, a free-standing editing domain homolog, ProXp-ala, is encoded in many organisms and has been shown to hydrolyze Ala-tRNA-Pro in trans. Here, we report that ProXp-ala from plants has a unique C-terminal extension domain that is missing from bacterial and higher eukaryotic ProXp-ala domains. To determine the function of the C-terminal extension in plant ProXp-ala, we designed and purified a truncated Arabidopsis thaliana ProXp-ala variant (At Delta;C-ProXp-ala). Circular dichroism spectroscopy showed that the secondary structure of ProXp-ala does not appear to be perturbed upon truncation. In vitro deacylation of Ala-tRNA-Proby both wild-type (WT) and At Delta;C-ProXp-ala showed that the truncation resulted in an 15-fold decrease in the deacylation rate. To establish whether this was primarily due to a binding defect, deacylation assays were performed with increasing concentrations of ProXp-ala. Based on the recovery of near WT levels of ProXp-ala deacylation with increasing amounts of the truncated construct, we conclude that the C-terminal domain in plant ProXp-ala has a critical role in binding the tRNA substrate. Future studies will focus on identifying the tRNA elements that are recognized by the unique C-terminal extension of plant ProXp-ala.
References:
1. Das, Mom, et al. "Distinct tRNA recognition strategies used by a homologous family of editing domains prevent mistranslation." Nucleic acids research 42.6 (2014): 3943-3953.
2. Vargas-Rodriguez, Oscar, and Karin Musier-Forsyth. "Exclusive use of trans-editing domains prevents proline mistranslation." Journal of Biological Chemistry 288.20 (2013): 14391-14399.
Keywords: trans-editing , aminoacyl-tRNA synthetase, ProXp-ala