Electrospray ionization (ESI) and Fourier transform mass spectrometry (FTMS) presents a unique platform for the investigation of ligand-nucleic acid interactions. In this direction, my thesis project is aimed at the development of new methods based on ESI-FTMS for studying the interactions between potential drug candidates and protein/nucleic acid targets of antibacterial and antiviral interest. Previously, we have applied ESI-FTMS to study the interactions of novel compounds with the Escherichia coli 16S ribosomal A-site.¹ These studies were spurred on by the unmet need for the development of a new class of antibiotics, which bind to the bacterial 16S ribosomal RNA but do not possess toxic side effects and that are not susceptible to bacterial resistance.

More recently, we examined the interactions of representative members of different classes of nucleic acid ligands with the separate domains (SL2-4) of the human immunodeficiency virus type 1 (HIV-1) packaging signal (Y-RNA ) and their complexes with the nucleocapsid (NC) domain of the Gag polyprotein.² In the life cycle of HIV-1, these protein-nucleic acid interactions mediate the essential functions of genome recognition, dimerization, and packaging and provide very attractive opportunities for the development of novel anti-retroviral strategies. Different tactics targeting key structures involved in these processes have been proposed to disrupt these crucial steps in the viral life cycle.^3,4  Our selection of small molecules constitutes typical ligand scaffolds that may provide valid templates for the development of new anti-retroviral agents. In an effort to explain the selective inhibitory effects induced by these ligands on the different NC-stemloop complexes, we explored the possibility of employing tandem mass spectrometry, such as SORI-CID, to map the actual binding sites of selected ligands on individual stemloop domains of Y-RNA.⁵

Current studies have focused on exploiting the high resolution of ESI-FTMS to screen combinatorial libraries and natural libraries of biodiverse proteins. Combinatorial chemistry provides a platform for synthesis of mixtures of large numbers of compounds, thereby greatly increasing the diversity of molecular architectures available for screening.⁶ Because each small molecule with a unique elemental composition carries an intrinsic mass label corresponding to its exact molecular mass, identifying closely related library members bound to a macromolecular target requires only a measurement of exact molecular mass. The target and potential ligands do not require further synthetic steps, such as radio-labeling or fluorescent tagging, which are necessary for other techniques. A similar approach can also be applied to natural libraries of peptides that offer new opportunities for identifying effective, specific inhibitors of protein-protein and protein-nucleic acid interactions. These libraries comprise both random and structured peptides encoded by natural genes of diverse genomes.⁷ This approach may help not only in understanding the implications of each identified interaction, but also in the development of effective drugs targeted to particular protein functions. Therefore, the ability of mass spectrometry for rapid determination of binding properties in a high-throughput, parallel fashion presents a unique model for drug discovery using multiple macromolecular targets against a collection of small-molecule ligands.

References:

1. Maddaford, S.P.; et al.; Bioorg. Med. Chem. Lett. 2004 14, 5987
2. Turner, K.B.; et al.; Nucleic Acids Res. 2006 34, 1305
3. Rice, W.G.; et al.; Nature 1993 361, 473
4. Skripkin, E.; et al.; J. Biol. Chem.  1996 271, 28812
5. Turner, K.B.; et al.; J. American Soc. Mass Spectrometry. (submitted)
6. Sannes-Lowery, K.A.; et al.; Inter. J. Mass Spec.  2004 238, 197
7. Watt, P.M.; Nature Biotech. 2006 24, 177