Molecular basis of plant secondary metabolites - proteins interaction

Abstract

The secondary metabolites found in plants represent an extremely rich source of novel chemical diversity for drug discovery and chemical biology programs.1–3 One of the main problems in drug discovery from plant small molecules is the identification of their molecular targets: many compounds have been found to be more promiscuous than originally anticipated, which can potentially lead to side effects, but which may also open up additional medical uses. The drug poly-pharmacological activity can be understood only if its interactions with cellular components are comprehensively characterized. Thus the identification of target proteins and investigation of ligand-receptor interactions represents an essential step in the process of plant drug discovery and development.4,5 However, nature is far more complex, and it is only with multidisciplinary collaborative research encompassing many disciplines that such targets can be successfully studied. Owing to our interest in the field of bioactive plant molecules, we have developed approaches to target identification based on chemical proteomics procedures, supported by spectroscopic and spectrometric data, Surface Plasmon Resonances (SPR) analyses and biochemical tests.5,6 Chemical proteomics is a powerful mass spectrometry-based affinity chromatography approach aimed to identify a set of proteins captured by a small molecules.6,7 The investigated molecule has to be anchored to a solid support through a flexible space arm, and then incubated with a lysate, from cell or tissues, to obtain the interaction between the immobilized compound (bait) and its protein targets. After several washing steps, proteins tightly bound to the beads are eluted and subjected to electrophoresis or gel free separation, followed by enzymatic digestion. The obtained peptide mixtures are then submitted to MS analyses and data base search for protein identification. This study allows the description of all potential macromolecular targets of a small bioactive molecules in a single experiment, leading to a complete and selective target mapping of a drug candidate. We have recently applied this approach to the study of cellular targets of some bioactive plant small molecules showing interesting protein targets; achieved results will be described in this communication.8,9 Koch, A.M., Schuffenhaue, A., Scheck, M., Wetzel, S., Casaulta M., Odermatt A., Ert P., Waldmann E., (2005) PNSA, 102:17272–17277. Clatdy, J., Walsh, C., (2004) Nature, 432:829–837. Li, J.W., Vederas, J.C. (2009) Scienze, 325: 161.166 Rix, U., Superti-Furga G., (2009) Nat. Chem. Biol., 5: 616–624. Bantscheff, M., Scholten, A., Heck. A. J., (2009) Drug Discov. Today14: 1021–1029. Dal Piaz, F.,Malafronte, N., Romano, A.,Gallotta, D., Belisario, M.A., Bifulco, G., Gualtieri, M.J., Sanogo, R., De Tommasi, N., Pisano C., (2012) Phytochemistry 75: 78–89 Dal Piaz, F.,Vassallo, A., Lepore, L., Tosco, A., Bader, A., De Tommasi, N., (2009)J. Med Chem, 52:3814–3828. Nigro, P., Dal Piaz, F., Gallotta, D., De Tommasi, N., Belisario, M. A., (2008) Free Rad Biol Med 45: 875–884. Tarallo, V., Lepore, L., Marcellini, M., Dal Piaz, F., Tudisco, L., Ponticelli, S., Lund, F.W.; Roepstoff., P., Orlandi, A., Pisano, C., De Tommasi, N., De Falco, S., (2011) JBC 286: 19641–19651.