Influence of 5-halogenation on the structure of protonated uridine: irmpd action spectroscopy and theoretical studies of the protonated 5-halouridines

The chemical and structural diversity and the extent of post-transcriptional modification of RNA is remarkable! Presently, there are 142 different naturally-occurring and many more synthetically modified nucleosides known. Uridine (Urd) is the most commonly modified nucleoside among those that occur naturally, but has also been an important target for synthesis and development of modified nucleosides for pharmaceutical applications. Indeed, modified nucleosides are of pharmaceutical interest due to their bioactivities. In particular, 5-bromouridine (br$^{5}$Urd) has been shown to exhibit antiviral activity to human immunodeficiency virus and has been used in RNA labeling studies. Halogenation is a common modification employed in pharmaceutical studies that enables systematic variation is the electronic properties of the molecule of interest due to the availability of halogen substituents that vary in size, dipole moment, polarizability, and electron withdrawing properties. In order to elucidate the influence of 5-halogenation on the intrinsic gas-phase structure and stability on the protonated form of Urd, synergistic spectroscopic and theoretical studies of the protonated forms of the 5-halouridines are performed here, where x$^{5}$Urd = 5-fluorouridine (f$^{5}$Urd), 5-chlorouridine (cl$^{5}$Urd), br$^{5}$Urd, and 5-iodouridine (i$^{5}$Urd). Infrared multiple photon dissociation (IRMPD) action spectra of the protonated forms of the 5-halouridines, [x$^{5}$Urd+H]$^{+}$, are measured over the IR fingerprint region using the FELIX free electron laser and the hydrogen stretching region using an OPO/OPA laser from 3300-3800 \wn. Complementary electronic structure calculations are performed to determine the stable low-energy conformations available to these species and to predict their IR spectra. Comparative analyses of the measured IRMPD spectra and predicted IR spectra are performed to elucidate the preferred sites of protonation, and the low-energy tautomeric conformations that are populated by electrospray ionization to be determined. Comparisons among these systems and to results previously reported for the protonated form of uridine, [Urd+H]$^{+}$, provides insight into the impact of the 5-halogen substituent on the structures and IR signatures