The RNA cleavage via internal transesterification is a fundamental reaction that is involved in RNA processing and metabolism, and the regulation thereof, but the mechanistic details have not been fully elucidated. Mechanistic studies will deepen our understanding not only of spontaneous RNA degradation under physiological conditions but also of ribozyme- and RNase-catalyzed RNA cleavage and splicing, and may facilitate the rational design of artificial ribonucleases for therapeutic RNA targeting.
The ribose moieties in RNA mainly exist in two puckered conformations, designated North (N) and South (S), that are in equilibrium; the former is thermally more stable and thus predominates. The authors introduced U6,3′‑Methano and U4’‑Me into oligonucleotides and confirmed that they were constrained in North and South conformation respectively by NMR. RNA cleavage rates were found to decrease in the order South-constrained ribonucleotide > native ribonucleotide ≫ North-constrained counterpart. DFT calculation results demonstrated that RNA cleavage via the S-type TS (Pathway II) is kinetically more favorable than via the N-type TS (Pathway I). Therefore, a rapid North to South conformational flip followed by internal transesterification via South transition states seems kinetically more favorable for RNA cleavage. The above-described results indicate that the ribose conformation plays an important role in modulating RNA cleavage via internal transesterification.
These results shed light on the effect of ribose conformation on RNA cleavage via internal transesterification and can be expected to improve our understanding of the mechanism of RNA cleavage catalyzed by RNases and ribozymes as well. In addition, the principle revealed may facilitate the rational design of catalysts for site-specific RNA cleavage and functionalized RNA molecules for biomedical applications.
Link: https://pubs.acs.org/doi/10.1021/jacs.8b06313.
