Poster abstracts

Poster number 24 submitted by Michael Ignatov

tRNA-dependent Switch in Editing Mechanisms Used by Class II ProRS

Michael Ignatov (Dept. of Chemistry, Ohio State University), Kathryn Splan (Dept. of Chemistry, University of Minnesota), Karin Musier-Forsyth (Depts. of Chemistry and Biochemistry, Ohio State University)

Abstract:
Aminoacyl-tRNA synthetases are essential enzymes, which charge amino acids to their cognate tRNAs in a two-step reaction that involves aminoacyl-adenylate formation followed by aminoacyl-tRNA synthesis. Unique amino acid recognition can generally not be achieved and some amino acids are mischarged by synthetases. To achieve a high degree of fidelity during protein synthesis, an editing mechanism is required to clear mischarged tRNAs. In the so-called double-sieve mechanism, amino acids similar in size and charge can be activated and transferred to the tRNA at the synthetic active site of the enzyme and are then transferred to the editing domain for hydrolysis of non-cognate aminoacyl-tRNA. Pre- and post-transfer editing pathways are responsible for clearing of non-cognate adenylates and aminoacyl-tRNAs, respectively. Here we present a study of the editing mechanisms used by ProRSs from all three Kingdoms of life to clear misactivated Ala-AMP and mischarged Ala-tRNA^Pro. In the presence of tRNA, post-transfer editing is the dominant pathway of editing. In contrast, in the absence of tRNA, pre-transfer editing of misactivated Ala-AMP occurs primarily through enzymatic hydrolysis at the synthetic active site. In the case of human ProRS, formation of Ala-AMP is rate limiting compared to its hydrolysis. In the case of E. coli and Methanococcus jannaschii ProRS, hydrolysis of the adenylate is the rate-limiting step.

Keywords: aminoacyl-tRNA synthetase, editing mechanism, ProRS