The modification of the wobble base of tRNA Glu modulates the translation rate of glutamic acid codons in vivo

Abstract

In Escherichia coli, uridine in the wobble position of tRNA Glu and tRNA Lys is modified to mnm 5s 2U34. This modification is believed to restrict the base-pairing capability, i.e. to prevent misreading of near-cognate codons and reduce the efficiency of cognate codon reading, especially of codons ending in G. We have determined the influence of the 5-methylaminomethyl and the 2-thio modifications of mnm 5s 2U34 in tRNA Glu on the translation rate of the glutamate codons GAA and GAG in vivo. In wild-type cells, GAG is translated slower (7.7 codons/second) and GAA faster (18 codons/second) than the average codon (13 codons/second). Surprisingly, tRNA Glu lacking the 5-methylaminomethyl group, thus containing s 2U34, translated GAA twofold faster (47 codons/second) and GAG fourfold slower (1.9 codons/second) than fully modified tRNA Glu. In contrast, tRNA Glu that contains mnm 5U34 instead of mnm 5s 2U34 translated GAA fourfold slower (4.5 codons/second) and GAG only 20% slower (6.2 codons/second). Clearly, the 5-methylaminomethyl group of mnm 5s 2U34 facilitates base-pairing with G while decreasing base-pairing with A, resulting in rates of translation of GAG and GAA that approach that of the average codon. The 2-thio group increases the recognition of GAA and has only a minor effect on the decoding of GAG. Furthermore, the 2-thio group is important for aminoacylation (see the accompanying paper). These data imply that the function of mnm 5s 2U34 may be different from what has been suggested previously.