Specific cleavages of pure tRNAs by plumbous ions

Abstract

After renaturation some pure tRNAs were submitted to the action of lead acetate 1 . 10(-3) M at pH 7.3 and at 37 degrees C in the presence of either 1 M or 0.5 M NaCl. These tRNAs were specifically cleaved by Pb2+. The exact cleavage points were determined by analysing the oligonucleotides obtained from three yeast tRNAs. In 1 M NaCl, tRNA(Phe) is cleaved after the hUp17 and partially cleaved after Cp73. In 0.5 M NaCl, there are cleaveages after hUp16, hUp17 as well as a partial one after pGP1. In 1 M NaCl tRNA(Asp) is not cleaved, whereas in 0.5 M NaCl 50% of the molecules are cleaved in the anticodon region after Up35, 14% after hUp19 and 6% after hUp16. In 1 M NaCl tRNA(Val) is cleaved in the hU loop: 40% after hUp16 and 60% after CP17. The action of lead on five other pure tRNAs was studied on the analytical scale only, by polyacrylamide gel electrophoresis. They could be classified into two familites, one cleaved mainly in the hU loop, the other in the anticodon loop. The minimal concentrations of Pb2+ required for cleavage were determined for several tRNAs, the most sensitive of which, yeast tRNA(Val), being still cleaved with a concentration of 5 . 10(-6) M in 0.15 M NaCl. Although the cleavage often occurs after hUp, poly (hU) is less sensitive than poly(U). This and other results indicate that cleavages depend more on the conformation than the sequence of the polynucleotide chain, bends in the tertiary structure being lead-sensitive sites. Finally, the amino acid acceptance activities of cleaved tRNA(Phe) and tRNA(Asp) were determined.