A sequence element that tunes Escherichia coli tRNA_GGC ^Ala to ensure accurate decoding

Mutating the rare A32-U38 nucleotide pair at the top of the anticodon loop of Escherichia coli tRNA_GGC^Ala to a more common U32-A38 pair results in a tRNA that performs almost normally on cognate codons but is unusually efficient in reading near-cognate codons. Pre-steady state kinetic measurements on E. coli ribosomes show that, unlike the wild-type tRNA _GGC^Ala, the misreading mutant tRNA_GGC ^Ala shows rapid GTP hydrolysis and no detectable proofreading on near-cognate codons. Similarly, tRNA_GGC^Ala mutated to contain C32-G38, a pair that is found in some bacterial tRNA_GGC ^Ala sequences, was able to decode only the cognate codons, whereas tRNA_GGC^Ala containing a more common C32-A38 pair was able to decode all cognate and near-cognate codons tested. We propose that many of the phylogenetically conserved sequence elements present in each tRNA have evolved to suppress translation of near-cognate codons.