Impact of 3-deazapurine nucleobases on RNA properties

Deazapurine nucleosides such as 3-deazaadenosine (c(3)A) are crucial for atomic mutagenesis studies of functional RNAs. They were the key for our current mechanistic understanding of ribosomal peptide bond formation and of phosphodiester cleavage in recently discovered small ribozymes, such as twister and pistol RNAs. Here, we present a comprehensive study on the impact of c(3)A and the thus far underinvestigated 3-deazaguanosine (c(3)G) on RNA properties. We found that these nucleosides can decrease thermodynamic stability of base pairing to a significant extent. The effects are much more pronounced for 3-deazapurine nucleosides compared to their constitutional isomers of 7-deazapurine nucleosides (c(7)G, c(7)A). We furthermore investigated base pair opening dynamics by solution NMR spectroscopy and revealed significantly enhanced imino proton exchange rates. Additionally, we solved the X-ray structure of a c(3)A-modified RNA and visualized the hydration pattern of the minor groove. Importantly, the characteristic water molecule that is hydrogen-bonded to the purine N3 atom and always observed in a natural double helix is lacking in the 3-deazapurine-modified counterpart. Both, the findings by NMR and X-ray crystallographic methods hence provide a rationale for the reduced pairing strength. Taken together, our comparative study is a first major step towards a comprehensive understanding of this important class of nucleoside modifications.

Automated summary

The study comprehensively investigates the impact of deazapurine nucleosides on RNA properties, showing that they can significantly decrease thermodynamic stability of base pairing and enhance base pair opening dynamics. The findings by NMR and X-ray crystallography provide a rationale for the reduced pairing strength, marking a major step towards a comprehensive understanding of this important class of nucleoside modifications.