Chemical Proteomics Partners identification of Cycloamphilectene

The identification of the cycloamphilectene cellular interactome has been performed, through chemical proteomics, to unambiguously define its mechanism of action and thus to understand its pharmacological effect. To achieve this goal, a small bioactive molecule (cycloamphilectene) linked to a matrix through a spacer arm “fishes out” its specific interactors from a cell lysate or a tissue extract. Once eluted, cellular partners are separated by SDS-PAGE and identified by high resolution MS and bioinformatics analysis . MS and MS/MS data were acquired using a Q-TOF Premier mass spectrometer (Waters Corp., Micromass, Manchester, United Kingdom). The five most intense ions were automatically chosen by the MassLynx (4.1) software and fragmented. After mass spectrometric measurements, data were automatically processed by ProteinLynx Global Server software to generate peak lists for protein identifications. Database searches were carried out with MASCOT server. The SwissProt human database (release 2010_11 of 02 Nov 10, 522019 sequences, 184241123 residues) was searched, allowing 2 missed cleavages, carbamidomethyl (C) as fixed modification o and oxidation (M) and phosphorylation (ST) as variable modifications. The peptide tolerance was set to 80 ppm and the MS/MS tolerance to 0.8 Da. The Raw data reported herein contains 4 different chemical proteomics experiments carried out to identify cycloamphilectene cellular partner.