A functional mutant of tRNA(2Arg) with ten extra nucleotides in its TFC arm

Abstract

An unusual, spontaneously arising mutant of Escherichia coli tRNA(2Arg) has been deduced to have two extra nucleotides in its anticodon loop and a duplication of ten nucleotide residues in the TFC loop. This conclusion is based on its gene sequence, Northern blot analysis of isolated tRNA and the size of the in vitro-processed tRNA product. In vitro analyses showed that the mutant precursor is processed normally, albeit inefficiently, to a mature tRNA species 12 nucleotides longer than the wild-type. In addition, the mature tRNA functions as a frameshift suppressor in vivo. Several related mutants with more conservative changes within the gene sequence were similarly shown to be accurately processed, albeit with varying degrees of efficiency less than that of the wild-type. These results indicate that in spite of the high degree of evolutionary conservation of the tertiary structure of tRNAs, and the fact that no such naturally-occurring variant has been found, a greatly enlarged tRNA is capable of functioning in protein synthesis. The data also indicate that recognition sites for correct processing of precursor tRNAs may be unexpectedly tolerant of unusual context, and may depend on some specific features of the tertiary structure rather than the overall structure for accurate processing.