An Innovative Method To Study Target Protein−Drug Interactions by Mass Spectrometry

Abstract

We report the combination of chemical cross-linking and high-resolution mass spectrometry for analyzing conformational changes in target proteins that are induced by drug binding. With this approach conformational changes in the peroxisome proliferator-activated receptor alpha (PPARalpha) upon binding of low-molecular weight compounds were readily detected, proving that the strategy provides a basis to efficiently characterize target protein-drug interactions.

Probing the Conformation of the ISWI ATPase Domain With Genetically Encoded Photoreactive Crosslinkers and Mass Spectrometry

“…With advances in MS instrumentation and computational analysis of the MS data, it became possible to precisely determine the residues involved in the crosslink (3)(4)(5). Crosslinks provide constraints on the through-space distance of the attachment sites, and this information can aid structure prediction (6,7), can distinguish protein conformations (8), and can identify the interaction surface between proteins (9,10) and between proteins and their ligands (11). Crosslinking-MS-based methods are widely applicable as they only require microgram quantities of protein, are not limited by protein size or solubility and are relatively tolerant against sample heterogeneity.…”

Mass spectrometry approaches to monitor protein–drug interactions

“…Changes in the solvent exposed surface of proteins are typically detected by hydrogen/ deuterium (H/D) exchange experiments [19], by altered accessibility of distinct amino acid side chains to chemical reactions [20], by cross-linking [21] or by probing the susceptibility to proteolytic degradation [22].…”

“…Recently, a combination of chemical cross-linking and high-resolution mass spectrometry has been reported for the analysis of drug-induced conformational changes in target proteins [33] (Fig 2b). The induced conformational changes of peroxisome proliferator-activated receptors (PPARs) upon binding of the antagonist (GW6471) or agonist (YS81) was investigated using amine-reactive homobifunctional cross linkers [21,34]. Drug-induced conformational changes led to altered spatial proximities of lysine side chains resulting in differential cross-link formation.…”

Flow injection-capillary electrophoresis frontal analysis method for the study of the interactions of a series of drugs with human serum albumin

“…A variety of methodologies such as fluorescence spectroscopy (FLU) [6], equilibrium dialysis (ED) [7,8], HPLC [9,10], flow injection chemiluminescence or sequential injection analysis [11,12], mass spectrometry (MS) [13] and CE [4,14] have been developed to evaluate drug-protein interactions. FLU is a simple method and has advantages such as high sensitivity and small sample requirement, but it can only be applied in a limited field.…”