Two distinct regions in Staphylococcus aureus GatCAB guarantee accurate tRNA recognition

In many prokaryotes the biosynthesis of the amide aminoacyl-tRNAs, Gln-tRNA^Gln and Asn-tRNA^Asn, proceeds by an indirect route in which mischarged Glu-tRNA^Gln or Asp-tRNA^Asn is amidated to the correct aminoacyl-tRNA catalyzed by a tRNA-dependent amidotransferase (AdT). Two types of AdTs exist: bacteria, archaea and organelles possess heterotrimeric GatCAB, while heterodimeric GatDE occurs exclusively in archaea. Bacterial GatCAB and GatDE recognize the first base pair of the acceptor stem and the D-loop of their tRNA substrates, while archaeal GatCAB recognizes the tertiary core of the tRNA, but not the first base pair. Here, we present the crystal structure of the full-length Staphylococcus aureus GatCAB. Its GatB tail domain possesses a conserved Lys rich motif that is situated close to the variable loop in a GatCAB:tRNA^Gln docking model. This motif is also conserved in the tail domain of archaeal GatCAB, suggesting this basic region may recognize the tRNA variable loop to discriminate Asp-tRNA^Asn from Asp-tRNA^Asp in archaea. Furthermore, we identified a 3₁₀ turn in GatB that permits the bacterial GatCAB to distinguish a U1–A72 base pair from a G1–C72 pair; the absence of this element in archaeal GatCAB enables the latter enzyme to recognize aminoacyl-tRNAs with G1–C72 base pairs.