Distinct ways of G:U recognition by conserved tRNA binding motifs.

Throughout three domains of life, alanyl-tRNA synthetases (AlaRSs) recognize a G3:U70 base pair in the acceptor stem of tRNA as the major identity determinant of tRNA The crystal structure of the archaeon AlaRS in complex with tRNA provided the basis for G3:U70 recognition with residues (Asp and Asn) that are conserved in the three domains [Naganuma M, et al. (2014) 510:507-511]. The recognition mode is unprecedented, with specific accommodation of the dyad asymmetry of the G:U wobble pair and exclusion of the dyad symmetry of a Watson-Crick pair. With this conserved mode, specificity is based more on "fit" than on direct recognition of specific atomic groups. Here, we show that, in contrast to the archaeal complex, the enzyme uses direct positive (energetically favorable) minor groove recognition of the unpaired 2-amino of G3 by Asp and repulsion of a competing base pair by Asn. Strikingly, mutations that disrupted positive recognition by the enzyme had little or no effect on G:U recognition by the human enzyme. Alternatively, AlaRS selects G:U without positive recognition and uses Asp instead to repel a competitor. Thus, the widely conserved Asp-plus-Asn architecture of AlaRSs can select G:U in a straightforward (bacteria) or two different unconventional (eukarya/archaea) ways. The adoption of different modes for recognition of a widely conserved G:U pair in alanine tRNAs suggests an early and insistent role for G:U in the development of the genetic code.