title: The mechanism of pre-transfer editing in yeast mitochondrial threonyl-tRNA synthetase.
abstract: Accurate translation of mRNA into protein is a fundamental biological process critical for maintaining normal cellular functions. To ensure translational fidelity, aminoacyl-tRNA synthetases (aaRSs) employ pre-transfer and post-transfer editing activities to hydrolyze misactivated and mischarged amino acids, respectively. Whereas post-transfer editing, which requires either a specialized domain in aaRS or a trans-protein factor, is well described, the mechanism of pre-transfer editing is less understood. Here, we show that yeast mitochondrial threonyl-tRNA synthetase (MST1), which lacks an editing domain, utilizes pre-transfer editing to discriminate against serine. MST1 misactivates serine and edits seryl adenylate (Ser-AMP) in a tRNA-independent manner. MST1 hydrolyzes 80% of misactivated Ser-AMP at a rate 4-fold higher than that for the cognate threonyl adenylate (Thr-AMP) while releasing 20% of Ser-AMP into the solution. To understand the mechanism of pre-transfer editing, we solved the crystal structure of MST1 complexed with an analog of Ser-AMP. The binding of the Ser-AMP analog to MST1 induces conformational changes in the aminoacylation active site, and it positions a potential hydrolytic water molecule more favorably for nucleophilic attack. In addition, inhibition results reveal that the Ser-AMP analog binds the active site 100-fold less tightly than the Thr-AMP analog. In conclusion, we propose that the plasticity of the aminoacylation site in MST1 allows binding of Ser-AMP and the appropriate positioning of the hydrolytic water molecule.
authors: Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520, USA.
authors: Jiqiang
authors: J
authors: Ling
authors: Kaitlyn M
authors: KM
authors: Peterson
authors: Ivana
authors: I
authors: Simonovic
authors: Dieter
authors: D
authors: Söll
authors: Miljan
authors: M
authors: Simonovic
mesh_terms: D018360:Crystallography, X-Ray; D008928:Mitochondria / Q000201:enzymology* / Q000235:genetics; D024101:Mitochondrial Proteins / Q000737:chemistry* / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017393:RNA Editing; D012331:RNA, Fungal; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics; D013914:Threonine-tRNA Ligase / Q000737:chemistry* / Q000235:genetics
chemical_list: D024101:Mitochondrial Proteins; D012331:RNA, Fungal; D029701:Saccharomyces cerevisiae Proteins; D013914:Threonine-tRNA Ligase