MManassantin A iTRAQ-SPROX Drug Mode-of-Action

ABSTRACT: Manassantin A is a natural product that has been isolated from the perennial herb Saururus chinensis Baill and the aquatic plant Saururus cernuus. Manassantin A has been shown to possess potent hypoxia inducible factor 1 alpha (HIF-1α) inhibitory activity in a cell-based assay screen of thousands of natural products. Manassantin A holds promise as an anti-cancer drug since it has been shown to selectively target tumor cells over normal cells. Due to the complex biological pathways involved in cancer and hypoxia, it is difficult to determine the mode-of-action by which manassantin A inhibits HIF-1. While some of the biological activities of manassantin A have been discovered in various cell-based activity assays, the molecular basis of manassantin A’s biological activities is not well characterized. The proteins in a hypoxic MDA-MB-231 cell lysate were screened for interactions with manassantin A using large scale experiments to uncover novel manassantin A interactions that lead to the drug’s HIF-1 inhibition and anti-cancer activity. Two energetics-based approaches were utilized in this manassantin A mode-of-action study: iTRAQ-SPROX and SILAC-Pulse Proteolysis. In these energetics-based approaches, protein stability is measured using the chemical denaturant dependence of either a methionine oxidation reaction (iTRAQ-SPROX) or a thermolysin protease digest (SILAC-Pulse Proteolysis). Using boh of these techniques, the stability of proteins in the absence and presence of excess manassantin A was monitored to assess ligand-induced protein stability changes.

INSTRUMENT(S): LTQ Orbitrap Elite

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Permanent Cell Line Cell, Cell Culture

DISEASE(S): Breast Cancer

SUBMITTER: Michelle Geer  

LAB HEAD: Michael C. Fitzgerald

PROVIDER: PXD002304 | Pride | 2016-07-12

REPOSITORIES: Pride