Mechanism of oligonucleotide hybridization with the 3 '-terminal region ofthe yeast tRNA(Phe)

Interaction of yeast phenylalanine tRNA with oligonucleotides complementary to its 3'-terminal nucleotide sequence was thoroughly studied. Using the g el retardation technique, thermodynamic and kinetic parameters of the tRNA complexation in physiological conditions were determined. Analysis of depen dences of the complex formation on the oligonucleotide concentration and in cubation time showed that this process proceeds in two stages. At the first stage, a metastable complex of the oligonucleotide with the open, single-s tranded sequence ACCA at the 3' end of tRNA rapidly forms. The second stage involves a slow intramolecular rearrangement of the resulting metastable c omplex into a full-sized heteroduplex accompanied by the tRNA(Phe) unfoldin g. The data gained suggest that the RNA unfolding stage is limiting in the interaction of oligonucleotides with natural RNAs. Principles of selection of optimal hybridization probes and antisense oligonucleotides are discusse d.