Nuclear magnetic resonance study of the codon-anticodon interaction in Bombyx mori tRNA(GCCGly)

NMR spectra of Bombyx mori tRNA(GCCGly) were recorded in the GCC absence and presence of the oligonucleotide, GGCUp, which contains the codon sequence, GGC. The difference between the spectra with and without the codon oligonucleotide indicates the appearance of five new imino proton peaks. For the assignment of these peaks, G*GCUp, in which the 5'-terminal G was enriched with 95% 15N, was prepared (G*, 15N-labeled guanosine). In the imino proton spectrum of B. mori tRNA(GCCGly) on the addition of G*GCUp, the peak at 12 ppm became a doublet due to coupling with 15N nuclei. In the two-dimensional 1H-15N heteronuclear multiple-quantum correlation (HMQC) spectrum, only the peak at 12 ppm was observed, and thus it was assigned to the imino proton of the 5'-terminal G of GGCUp interacting with tRNA(GCCGly). Judging from the temperature effect and chemical shifts, the five new imino proton peaks are presumed to be due to three G.C base pairs, induced by the codon-anticodon interaction, and one U.U base pair, induced by an interaction between the 3' terminal U of GGCUp and U33 neighboring the anticodon. The binding of three trinucleotides (GGCp, GGUp and GCUp) to B. mori tRNA(GCCGly) was also investigated. Ultracentrifugation analysis showed that tRNA(GCCGly) underwent dimerization through the anticodon-anticodon interaction, but the dimerization was broken on addition of GGCUp. On 1H-NMR and ultracentrifugation analysis, it was found that GCUp not complementary to the anticodon also binds to the anticodon loop.