The first example of sequence-specific non-uniformly 13C5 labelled RNA: Synthesis of the 29mer HIV-1 TAR RNA with 13C Relaxation Window

Abstract

A complete synthetic protocol as well as 1H- and 13C-NMR data on the monomer building blocks used for the solid-phase synthesis of specifically 13C-labelled (99 atom % 13C) stem ( 27A and 43G), bulge ( 24C) and loop ( 31U) regions of 29mer HIV-1 TAR RNA hairpin starting from the 13C 6- D -glucose are presented. The complex NMR spectra of 13C-labelled monomer building blocks, due to the interaction of various 13C and 1H spins, have been assigned. It has been demonstrated by heteronuclear 2D NMR that the non-uniform labelling of the HIV-1 TAR 29mer RNA achieved herein by chemical synthesis provides an optimal opportunity to perform full T 1 and T 2 relaxation measurements (the “NMR Relaxation Window”) of each type of sugar-carbons for all four strategically placed 13C-labelled residues in a unique and unprecedented manner because of minimal overlap of 13C resonances compared to uniformly labelled oligo-RNA.