Glycine transfer RNA of Escherichia coli: II. Impaired GGA-recognition in strains containing a genetically altered transfer RNA; Reversal by a secondary suppressor mutation

Abstract

The glycine-specific tRNA from Escherichia coli is separable into three components (tRNA I Gly, tRNA II Gly and tRNA III Gly) by chromatography over BD-ellulose. ‡ ‡ Abbreviations used: BD-cellulose, benzoylated DEAE-cellulose; genetic symbols are explained in the legend of Table 1. Triplet binding studies reveal that tRNA I Gly recognizes predominantly GpGpG, tRNA II Gly recognizes both GpGpG and GpGpA, and tRNA III Gly recognizes GpGpU and GpGpC. Strains carrying a suppressor ( glyTsu) of the trpA36 mutation (Gly → Arg, GGA → AGA) lack a functional tRNA II Gly and, as a result, exhibit severe pleiotropic effects. As expected, the binding of [ 14C]glycyl-tRNA from glyTsu strains to ribosomes is only weakly stimulated by GpGpA (8% of the control), while binding in the presence of GpGpG, GpGpU and GpGpC is relatively normal. On the other hand, suppressed strains ( glyUsu) lacking a functional tRNA I Gly do not show these adverse effects, and tRNA from these strains displays a nearly normal codon recognition pattern. The pleiotropic effects seen in glyTsu strains lacking a normal tRNA II Gly are reversed either by (a) the presence of a normal glyT + gene in the same organism, or (b) by secondary mutations ( ins) unlinked to the glyT region. The GpGpA-stimulated binding of glyTsu ins tRNA to ribosomes is partially restored to normal. Transfer RNA from ins strains contains a new tRNA Gly species (tRNA III Gly), easily detectable by BD-cellulose chromatography. This new tRNA III Gly, binds to ribosomes in the presence of GpGpA or GpGpG, but not GpGpU or GpGpC. The results indicate that tRNA III Gly is formed by a genetically induced alteration of a portion of tRNA III Gly. It seems likely that the ins mutation results in a GGU/C → GGA/G change in a redundant tRNA III Gly species, thus relieving the pleiotropy induced by the loss of a normal tRNA II Gly.