Reduced misreading of asparagine codons by Escherichia coli tRNALys with hypomodified derivatives of 5-methylaminomethyl-2-thiouridine in the wobble position

It has been suggested that modified nucleosides of the xm(5)(s(2))U(m)34-type restrict the wobble capacity of the base, and that their function is to prevent misreading in the third position of the codon in mixed codon family boxes that encode two different amino acids. In this study in Escherichia coli, the misreading in vivo of asparagine codons in bacteriophage MS2 mRNA by different hypomodified derivatives of tRNA(Lys), normally containing 5-methylaminomethyl-2-thiouridine (mnm(5)s(2)U34) in the wobble position, has been analysed. Contrary to what would be predicted from the general hypothesis for the function of mnm(5)s(2)U, it was found that the misreading of asparagine codons by tRNA(Lys) was greatly reduced in the mnmA (formerly asuE or trmU) and mnmE (formerly trmE) mutants which contain the hypomodified mnm(5)U34 and s(2)U34, respectively, instead of the fully modified mnm5s2U34. In addition, it was found that these hypomodified tRNAs were efficiently charged with lysine in vivo, under the growth conditions employed. The latter result is at variance with results obtained in vitro. The results are discussed in relation to the postulated function for modified nucleosides of the xm(5)s(2)U type. (C) 1998 Academic Press.