 |
Journal articles
Monitoring ligand modulation of protein-protein interactions by mass spectrometry: estrogen receptor alpha-SRC1.
Abstract : Many drugs and chemicals exert their biological effect by modulating protein-protein interactions. In vitro approaches to characterize these mechanisms are often based on indirect measurements (e.g., fluorescence). Here, we used mass spectrometry (MS) to directly monitor the effect of small-molecule ligands on the binding of a coactivator peptide (SRC1) by the human estrogen receptor alpha ligand binding domain (hERalpha LBD). Nanoelectrospray mass spectrometry (nanoESI-MS) and high-mass matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) combined with chemical cross-linking were employed to follow these processes. The chemical cross-linking protocol used prior to high-mass MALDI analysis allows detection of intact noncovalent complexes. The binding of intact hERalpha LBD homodimer with two coactivator peptides was detected with nanoESI-MS and high-mass MALDI-MS only in the presence of an agonist ligand. Furthermore, high-mass MALDI-MS revealed an increase of the homodimer abundance after incubating the receptor with a ligand, independent of the ligand character (i.e., agonist, antagonist). The binding characteristics of the compounds tested by MS correlate very well with their biological activity reported by cell-based assays. High-mass MALDI appears to be an efficient and simple tool for directly monitoring ligand regulation mechanisms involved in protein-protein interactions. Furthermore, the combination of both MS methods allows identifying and characterizing endocrine-disrupting compounds or new drug compounds in an efficient way.
https://www.hal.inserm.fr/inserm-00350740
Contributor : Maité Peney <>
Submitted on : Wednesday, January 7, 2009 - 3:47:21 PM
Last modification on : Monday, December 14, 2020 - 3:48:03 PM
Contributor : Maité Peney <>
Submitted on : Wednesday, January 7, 2009 - 3:47:21 PM
Last modification on : Monday, December 14, 2020 - 3:48:03 PM
Links full text
