Identification of transfer RNA suppressors in Escherichia coli: I. Amber suppressor su+2, an anticodon mutant of tRNA 2Gln

Abstract

In order to isolate the gene for amber suppressor su +2 ( SupE) in Escherichia coli, a non-defective su +2-transducing phage lambda was isolated in three steps: first, deletion derivatives of F′ su +2 gal (λ) were selected, linking su +2 to the right-hand prophage attachment site, att λ PB′; second, these F′-factors were relysogenized by λ and defective transducing phages, λd su +2, were produced by induction; and third, non-defective λp su +2 transducing phages were produced by recombination of λd su +2 isolates with λ. Upon infection by λp su +2, the production of transferRNAs accepting glutamine and methionine was markedly stimulated. Fingerprint analysis of these tRNAs revealed that they consisted of normal tRNA 2 Gln, mutant tRNA 2 Gln and tRNA m Met. The mutant tRNA 2 Gln carried a singlebase alteration from G to A at the 3′-end of the anticodon. The production of tRNA 1 Gln was not stimulated by the infection of λp su +2. We conclude that the wild-type allele of su +2 ( SupE) is the structural gene for tRNA 2 Gln, and the su +2 amber suppressor was derived by a single base mutation, changing the anticodon from CUG to CUA, in one of the multi-copy genes for tRNA 2 Gln. The fact that λp su +2 also induces the production of tRNA m Met suggests that this tRNA is encoded in the same chromosomal region of E. coli as is tRNA 2 Gln.