Year: 2019
Type: article
Abstract: The universally conserved N6-threonylcarbamoyladenosine (t6A) modification of tRNA is essential for translational fidelity. In bacteria, t6A biosynthesis starts with the TsaC/TsaC2-catalyzed synthesis of the intermediate threonylcarbamoyl adenylate (TC-AMP), followed by transfer of the threonylcarbamoyl (TC) moiety to adenine-37 of tRNA by the TC-transfer complex comprised of TsaB, TsaD and TsaE subunits and possessing an ATPase activity required for multi-turnover of the t6A cycle. We report a 2.5-Å crystal structure of the T. maritima TC-transfer complex (TmTsaB2D2E2) bound to Mg2+-ATP in the ATPase site, and substrate analog carboxy-AMP in the TC-transfer site. Site directed mutagenesis results show that residues in the conserved Switch I and Switch II motifs of TsaE mediate the ATP hydrolysis-driven reactivation/reset step of the t6A cycle. Further, SAXS analysis of the TmTsaB2D2-tRNA complex in solution reveals bound tRNA lodged in the TsaE binding cavity, confirming our previous biochemical data. Based on the crystal structure and molecular docking of TC-AMP and adenine-37 in the TC-transfer site, we propose a model for the mechanism of TC transfer by this universal biosynthetic system.
Source: Nucleic Acids Research
DOI: 10.1093/nar/gkz439
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $3,992
Awards: G1229526393, G1286251239, G1804828992, G1831385526, G2371548206, G262532246, G303061748, G315621713, G3344126041, G3488436650, G3629260272, G4235786860, G4280418065, G4581610328, RNA modification: Mechanism and links to other metabolic pathways, G495301720, G498139845, G5570207115, G6550344761, G7283024673, G8382000481, A Synchrotron Radiation Structural Biology Resource, G8925327636, G911061179, G969889393
OpenAlex ID: W2946423216