Phosphorolysis of the tRNA. Conformations of specific tRNAs and effect of the localized regions on the stability of the structure.

Abstract

Phosphorolysis studies of several specific tRNAs from Escherichia coli and yeast confirm the phenomenon observed with total tRNA: each tRNA can be differentiated into two classes of conformations, according to their ability (S class) or inability (R class) to be phosphorolyzed by polynucleotide phosphorylase. The proportion of these conformations at a given temperature varies from one specific tRNA to another. When the extent of phosphorolysis of different specific tRNAs are compared with the variation of hyperchromicity at different temperatures, we find that these two parameters are correlated. The phosphorolysis behavior of tRNA differs when it is phosphorolyzed in a mixture or alone. Studies with (E. coli) tRNATyr show that, at low temperature, the extent of phosphorolysis of this tRNA, assayed alone, is much higher than that of the total E. coli tRNA or other specific tRNAs tried; moreover, when this tRNA is mixed with (E. coli) tRNASer or included in the bulk of E. coli tRNA, it is scarcely attacked. In contrast, at high temperatures where all tRNA is completely degraded by the enzyme, there is no longer any difference in the rate of phosphorolysis between tRNATyr and tRNASer from E. coli. The differential phosphorolysis of these tRNAs at low temperatures is interpreted as a differential affinity of these substrates for the enzyme. One of the unique structural features of (E. coli) tRNATyr is the long nucleotide sequence which forms a long “extra” arm in the clover-leaf model. It is postulated that the presence of a long “extra” arm may be the reason for the observed low thermal transition at the first melting region of this tRNA and for the decreased affinity of the tRNA for the enzyme. Studies with yeast tRNASer, tRNAPhe, and bulk tRNA lead to similar results. The differences in the extent of phosphorolysis are discussed in connection with the topology of the tRNA structure, according to the length of the dihydrouridine loop and of its base-paired portion. It appears that tRNAs with a long dihydrouridine loop, a short base-paired portion and a long “extra” arm are best phosphorolyzed; those with a short dihydrouridine loop and a long base-paired region are more resistant.