Synthesis of Oligodeoxyribonucleotide Ethyl Phosphotriesters and Their Specific Complex Formation with Transfer Ribonucleic Acid

Oligodeoxyribonucleotide ethyl phosphotriesters, d-Tp(Et)Gp(Et)G and d-Tp(Et)Tp(Et)Cp(Et)A, were synthesized by a stepwise, chemical procedure. These triesters are complementary, respectively, to 3′-CpCpA-OH (the 3′-amino acid accepting terminus) and to -UpGpApA- (the anticodon region) of phenylalanine tRNA from yeast and Escherichia coli. Tritium-labeled triesters were prepared by exchange of the H-8 protons of adenine and guanine in the oligomers in tritiated water. The association constants for binding of the triesters to their complementary regions on tRNA were measured by equilibrium dialysis and were compared with those of oligodeoxyribonucleotides and oligoribonucleotides of the same sequences. In 1 M NaCl-10 mM MgCl₂ at 0°, the association constants of the oligomers with both tRNA^phe _yeast and tRNA^phe _coli are very similar. The association constants of the ribooligonucleotides are 8 to 20 times larger than those of the corresponding deoxyribooligonucleotides, while the deoxyribooligonucleotide triesters exhibit binding constants slightly higher than those of the deoxyribooligonucleotides. These differences are discussed in terms of the differences in conformations of the various oligomers. At low salt concentration (0.1 M NaCl, 1 mM EDTA), the oligonucleotide triesters have the same binding constants as at high salt concentration, whereas the corresponding deoxyribo- and ribooligonucleotides show a four- to sixfold decrease in their binding constants. This reflects the removal of the charge repulsion between the neutral triesters and the tRNA. The binding of oligomers to modified tRNA^phe _yeast was also examined. Removal of the Y base decreased the binding of anticodon-complementary oligomers sixfold while removal of the 3′-CpA residues decreased the binding of the 3′-CpCpA-OH complementary oligomers 6- to 20-fold. This study provides the chemical and physicochemical basis for the investigation of the biochemical effects of these triesters on the aminoacylation of tRNA which is reported in the following paper (Barrett, J. C., Miller, P. S., and Ts'o, P. O. P. (1974), Biochemistry 13, 4897).