Structure ofSaccharomyces cerevisiaemitochondrial Qri7 in complex with AMP

Abstract

N6-Threonylcarbamoyladenosine (t6A) is a modified tRNA base required for accuracy in translation. Qri7 is localized in yeast mitochondria and is involved in t6A biosynthesis. In t6A biosynthesis, threonylcarbamoyl-adenylate (TCA) is synthesized from threonine, bicarbonate and ATP, and the threonyl-carbamoyl group is transferred to adenine 37 of tRNA by Qri7. Qri7 alone is sufficient to catalyze the second step of the reaction, whereas the Qri7 homologues YgjD (in bacteria) and Kae1 (in archaea and eukaryotes) function as parts of multi-protein complexes. In this study, the crystal structure of Qri7 complexed with AMP (a part of TCA) has been determined at 2.94 Å resolution in a new crystal form. The manner of AMP recognition is similar, with some minor variations, among the Qri7/Kae1/YgjD family proteins. The previously reported dimer formation was also observed in this new crystal form. Furthermore, a comparison with the structure of TobZ, which catalyzes a similar reaction to t6A biosynthesis, revealed the presence of a flexible loop that may be involved in tRNA binding.

The structural and functional workings of KEOPS

“…developed a non-hydrolysable TC-AMP analogue (termed BK951) and reported its structure bound to the E. coli TsaD-TsaB heterodimer ( 84 ). The structure revealed that the adenine base portion of TC-AMP is ‘sandwiched’ between subdomain 2 and Kae1 specific insert I, similar to the nucleotide binding mechanism of Kae1 enzymes reported in previous structures ( 8 , 84 , 91 , 95 , 105 ). A cation bound by the metal binding triad serves to coordinate the carboxylate group of the threonine moiety of TC-AMP (Figure 3B ).…”

The ATP-mediated formation of the YgjD–YeaZ–YjeE complex is required for the biosynthesis of tRNA t6A in Escherichia coli

“…Interestingly, the yeast mitochondrial YgjD/Kae1 paralog, Qri7, is capable of the in vitro biosynthesis of tRNA t 6 A on its own if provided with TCA ( 12 , 22 ). The active site of YgjD/Kae1/Qri7 is characterized by a conserved metal-cluster that is contacting the phosphate moieties of a bound ATP ( 23 – 25 ). Mutations of this metal-binding motif abolish the biosynthesis of t 6 A ( 12 , 13 , 18 ).…”

Diversity of the biosynthesis pathway for threonylcarbamoyladenosine (t6A), a universal modification of tRNA

“…2), 30,31 YrdC and YgjD failed to produce t 6 A in vitro with transcript or t 6 A-deficient tRNA purified from yeast sua5D, 30 suggesting that the biosynthetic machinery for t 6 A biosynthesis required more than these 2 proteins. Over the last 2 years, a flurry of papers have reported the identification of these missing proteins, and elucidated the complete bacterial, 28,33 eukaryotic/archaeal, 29,40-42 and mitochondrial 32,43,44 biosynthetic pathways to t 6 A.…”