Catalytic and regulatory basis of tRNA t<sup>6</sup>A modification by the KEOPS complex.

Zhou, Li; Zhang, Zelin; Jin, Mengqi; Xie, Dengmiao; Wen, Han; Westhof, Eric; Lei, Dongsheng; Zhang, Wenhua · Nat Commun · 2026

basic_science · Level V

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Abstract

N<sup>6</sup>-threonylcarbamoyladenosine (t<sup>6</sup>A) at tRNA position 37 is essential for translational fidelity and cellular homeostasis. The multi-subunit KEOPS complex catalyzes t<sup>6</sup>A formation in Archaea and Eukarya. Here we present cryo-EM structures of C. elegans KEOPS in its apo and tRNA-bound states. tRNA binding induces concerted conformational rearrangements, distorting the anticodon loop to project A37 into the Kae1 active site. Kae1 recognizes the conserved G10-C25 pair and 36-UAA-38 motif. Bud32 directly contacts the anticodon and acceptor arms, coupling ATP hydrolysis to t<sup>6</sup>A catalysis in the distant Kae1 active site through long-range conformational changes. Cgi121 enhances catalytic efficiency through cooperative binding with Bud32 and the tRNA 3' CCA. Pcc1 stabilizes the anticodon loop and mediates KEOPS dimerization, enhancing tRNA binding and t<sup>6</sup>A activity. GAMOS-associated mutations cluster at functional hotspots within the KEOPS-tRNA complex. This study provides a structural framework for understanding KEOPS mechanism and its cellular roles.