Structural studies on phenylalanine transfer ribonucleic acid from yeast with the spectroscopic label formycin

Formycinmonophosphate (FMP) has been incorporated in to the 3′‐terminus of phenylalanine tRNA from yeast by tRNA nucleotidyl transferase. tRNAPhepCpCpF can be amino‐acylated with the cognate synthetase exhibiting the same Km and an about 50‐times lowered V compared to tRNAPhepCpCpA. Quantum yield of FMP fluorescence in tRNAPhepCpCpF is about four times lower than in the monomer. Comparative studies of tRNAPhepCpCpF and the oligonucleotide CpApCpCpF show that the differences between the fluorescence properties of tRNAPhepCpCpF and the monomeric formycin‐nucleotide are due only to stacking interactions and do not arise from the tertiary structure of tRNA. After periodate oxidation of the 3′‐terminal ribose of tRNAPhepCpCpF the full fluorescence of formycin is reconstituted. This emphasises the important role of the intact ribose ring for stabilization of stacking units. Periodate oxidation of tRNAPhepCpCpF followed by borohydride reduction leads to a tRNA that acts as competitive inhibitor in the aminoacylation reaction. This, together with its high quantum yield makes it a valuable tool in fluorescence spectroscopic studies of the aminoacylation reaction.