Importance of formylability and anticodon stem sequence to give a tRNA(Met) an initiator identity in Escherichia coli

In bacteria, the free amino group of the methionylated initiator tRNA is specifically modified by the addition of a formyl group. The functional relevance of such a formylation for the initiation of translation is not yet precisely understood. Advantage was taken here of the availability of the fmt gene, encoding the Escherichia coli Met-tRNA(f)/(Met) formyltransferase, to measure the influence of variations in the level of formyltransferase activity on the involvement of various mutant tRNA(f)/(Met) and tRNA(m)/(Met) species in either initiation or elongation in vivo. The data obtained established that formylation plays a dual role, firstly, by dictating tRNA(f)/(Met) to engage in the initiation of translation, and secondly, by preventing the misappropriation of this tRNA by the elongation apparatus. The importance of formylation in the initiator identity of tRNA(f)/(Met) was further shown by the demonstration that elongator tRNA(m)/(Met) may be used in initiation and no longer in elongation, provided that it is mutated into a formylatable species and is given the three G · C base pairs characteristic of the anticodon stem of initiator tRNAs.