RNA is arguably the most functionally diverse
biological macromolecule. In some cases a single discrete RNA sequence performs multiple roles, and this can be conferred by a complex three-dimensional structure. Such multifunctionality can also be driven or enhanced by the ability of a given
RNA to assume different conformational (and therefore functional) states. Despite its
biological importance, a detailed structural understanding of the paradigm of
RNA structure-driven multifunctionality is lacking. To address this gap it is useful to study examples from single-stranded positive-sense RNA viruses, a prototype being the
tRNA-like structure (TLS) found at the 3' end of the turnip yellow mosaic virus (TYMV). This TLS not only acts like a
tRNA to drive aminoacylation of the viral genomic (g)
RNA, but also interacts with other structures in the
3' untranslated region of the gRNA, contains the promoter for negative-strand synthesis, and influences several
infection-critical processes. TLS
RNA can provide a glimpse into the structural basis of
RNA multifunctionality and plasticity, but for decades its high-resolution structure has remained elusive. Here we present the crystal structure of the complete TYMV TLS to 2.0 Å resolution. Globally, the
RNA adopts a shape that mimics
tRNA, but it uses a very different set of intramolecular interactions to achieve this shape. These interactions also allow the TLS to readily switch conformations. In addition, the TLS structure is 'two faced': one face closely mimics
tRNA and drives aminoacylation, the other face diverges from
tRNA and enables additional functionality. The TLS is thus structured to perform several functions and interact with diverse binding partners, and we demonstrate its ability to specifically bind to ribosomes.