Structural and functional insights into tRNA recognition by human tRNA guanine transglycosylase

Eukaryotic tRNA guanine transglycosylase (TGT) is an RNA-modifying enzyme which catalyzes the base exchange of the genetically encoded guanine 34 of tRNAsAsp,Asn,His,Tyr for queuine, a hypermodified 7-deazaguanine derivative. Eukaryotic TGT is a heterodimer comprised of a catalytic and a non-catalytic subunit. While binding of the tRNA anticodon loop to the active site is structurally well understood, the contribution of the non-catalytic subunit to tRNA binding remained enigmatic, as no complex structure with a complete tRNA was available. Here, we report a cryo-EM structure of eukaryotic TGT in complex with a complete tRNA, revealing the crucial role of the non-catalytic subunit in tRNA binding. We decipher the functional significance of these additional tRNA-binding sites, analyze solution state conformation, flexibility, and disorder of apo TGT, and examine conformational transitions upon tRNA binding. [Display omitted] •Cryo-EM structure of tRNA-guanine transglycosylase and bound tRNA at 2.8 Å resolution•tRNA-binding sites of the non-catalytic subunit increase complex stability and affinity•SAXS analysis of apo TGT and TGT·tRNA complex reveals an extended conformation Sievers et al. determined the cryo-EM structure of human tRNA guanine transglycosylase in complex with tRNAAsp and measured the enzymatic activity and binding affinities with various RNA constructs and TGT mutants. Together, these experiments reveal the basis of tRNA recognition by TGT.