Sponsor
National Institutes of Health GM110588 to M.A.S. and D.I.-R., a grant from the Singapore-MIT Alliance for Research and Technology under the Antimicrobial Resistance Interdisciplinary Research Group to P.C.D., C.-K.C. is supported by a Human Frontier Science Program Cross-Disciplinary Fellowship. A portion of this research was supported by NIH grant R24GM154185 and performed at the Pacific Northwest Center for Cryo-EM (PNCC) with assistance from Marzia Miletto. Funding to pay the Open Access publication charges for this article was provided by NIGMS grant GM110588.
Published In
Nucleic Acids Research
Document Type
Article
Publication Date
6-24-2026
Subjects
Anticodon Loop Remodeling -- Chemistry
Abstract
N6-threonylcarbamoyladenosine (t6A) is a universal transfer RNA (tRNA) modification essential for translational fidelity. The modification is installed at position 37 of ANN-decoding tRNAs (N is A, U, G, or C) by transfer of a threonylcarbamoyl moiety from a pathway intermediate, catalyzed in bacteria by the TsaBD complex. Despite the strict requirement for the 36-UAA-38 sequence in substrate tRNAs, the structural basis for this specificity has remained unclear. We determined cryo-EM structures of the Thermotoga maritima t 6A synthase bound to unmodified and to natively modified tRNA carrying the t 6A37 modification, at a nominal resolution of 3.2 A˚ . In both structures, A37 is positioned in the active site, and the anticodon loop is remodeled into a zig-zag conformation not previously observed in tRNA, stabilized by conserved interactions with the RNA backbone at the TsaD/TsaB interface. The wobble and middle anticodon bases occupy shallow surface pockets without base-specific contacts, explaining tolerance to base identity at these positions. In contrast, U36 and A38 are recognized indirectly through formation of a base triple with U32. Additional D-stem contacts provide a second mode of indirect readout. Together with mutagenesis data, the structures reveal the molecular basis for restriction of t 6A37 to ANN-decoding tRNAs and the requirement for A38.
Rights
Copyright (c) 2026 The Authors Creative Commons License This work is licensed under a Creative Commons Attribution 4.0 International License.
Locate the Document
DOI
10.1093/nar/gkag599
Persistent Identifier
https://archives.pdx.edu/ds/psu/45208
Citation Details
Kutchuashvili, A., Scheleen, E., Guynes, G., Wu, J., Chan, C.-K., Dedon, P., Iwata-Reuyl, D., & Swairjo, M. A. (2026). Anticodon loop remodeling and D-stem shape drive the specific recognition of ANN-decoding tRNAs for t6A modification. Nucleic Acids Research, 54(11).