The structure of the anticodon loop of tRNAPhe from yeast as deduced from spectroscopic studies on oligonucleotides.

Abstract

AbstractThe hexanucleotide Gm‐A‐A‐Y‐A‐ψp excised from the anticodon loop of yeast tRNAPhe and its constituent oligonucleotides have been studied by ultraviolet absorption spectroscopy, static fluorescence, and circular dichroism. Gm‐Ap has a melting point of 45°C and a high melting enthalpy when compared with G‐Ap; hence 2′‐O‐methylation seems to stabilize stacking interactions. The nucleobase Y adjacent to the 3′‐side of the anticodon triplet interacts stronger with its 3′‐neighboring A than with its 5′‐neighboring A. It is concluded that the base Y disconnects the stack of the anticodon itself from the stack of the anticodon stem, thereby setting a reading frame for the mRNA in the course of protein biosynthesis. From the opposite signs of the short‐wavelength Cotton effects in the spectra of Gm‐A‐A‐Y‐Ap and Gm‐A‐A‐Y, it is concluded that Y after removal of its 3′ neighbor undergoes a dramatic change in its conformation. The fluorescence of the nucleobase Y upon addition of Mg2+ is enhanced in oligonucleotides longer than two. An identical enhancement is observed for tRNAPhe, indicating that this Mg2+ effect is a property of an oligonucleotide segment and does not reflect conformational changes of the whole tRNA. The data presented here reveal that the basic structural features of the anticodon loop are already present in the hexanucleotide Gm‐A‐A‐Y‐A‐ψp and are not determined by the overall structure of tRNA.