Valylation of tRNA-like transcripts from cloned cDNA of turnip yellow mosaic virus RNA demonstrate that the L-shaped region at the 3' end of the viral RNA is not sufficient for optimal aminoacylation.

Clones containing different lengths of cDNA corresponding to the 3' region of turnip yellow mosaic virus RNA were constructed and transcribed in vitro into the corresponding RNAs. Each transcript contained the L-shaped tRNA domain (N = 82) plus (i) in the case of 3 upstream sequences up to N = 93, 109 and 258; and (ii) in all cases an additional 6 nucleotide-stretch at the 5' end derived from the T7 promoter. The valylation of these molecules, as well as that of a fragment (N = 159) purified from viral RNA, was studied. Although all transcripts could be valylated by wheat germ valyl-tRNA synthetase, the 3 shorter fragments showed incomplete charging and slower rates, due mainly to lower Vmax values. Thus, although the tRNA-like L-shaped structure is the functional core permitting amino-acylation, upstream nucleotides between positions 82 and 159 play an important role in allowing the highest rates and levels of valylation. Structural arguments supporting this view are discussed.