Vibrational spectroscopies, NMR, UV absorption and molecular modeling by DFT calculations to elucidate RNA structures

Abstract

Ribo-nucleic acid (RNA) is an important biomolecule with a wide range of functions: transfer of the genetic code and genomic regulation, protein synthesis as well as catalytic properties [1]. From the structural point of view, RNA hairpins, which consist of an intramolecular double-helix (stem) capped by a certain number of unpaired nucleotides (loop), allow this molecule to fold back onto itself giving rise to its particular secondary and tertiary structures. In ribosomal RNAs, the majority of the loops include four ribonucleotides (tetraloops). It has been shown that over 70% of these tetraloops belong to GNRA and UNCG families (N = U, A, C., G and R = G, A) [2]. Furthermore, one can recall some of the biological functions of GNRA and UNCG tetraloops: UUCG is the terminal site of the reverse-transcriptase action, whereas GAAA participates in long-range tertiary interactions in catalytic RNAs. In addition, it has been shown that UNCG and GNRA tetraloops, owing to their particular structural features, are unusually stable (revealed by their high T_m values derived from the UV absorption melting files), provided that their stems include at least two G·C base-pairs, preferentially with a C·G one closing the tetraloop [3]. Thus, short hairpins with a comparable number of ribonucleotides in the loop and in the stem can easily be formed in aqueous phase. This is extremely important for spectroscopic techniques (NMR and vibrational spectroscopies) used to probe the structural features of these systems, because the signals arising from the loop ribonucleotides are not completely hidden by those arising from the stem.