Electron paramagnetic resonance characterization of drug‐binding to monoacylglycerol lipase using the dynamic signature of a protein spin‐label (779.3)

Abstract

In recent years, monoacylglycerol lipase (MGL) has been an important pharmaceutical target because of its relationship to pathways that have been implicated in cancer proliferation. The purpose of this study is to find an efficient and streamlined way of characterizing drug‐binding to MGL using electron paramagnetic resonance (EPR). The free nitroxide radical methyl thiosulfonyl spin label (MTSL) was used to label the single cysteine mutant S208C on a domain of MGL that is thought to constitute a “lid” covering the substrate binding pocket of MGL. EPR spectra shows reproducible line shape changes reflecting a significant reduction in the mobility of the lid when MGL binds reversible inhibitors. The results are consistent with NMR data that reveal an equilibrium between the putative “open” and “closed” positions of the lid as a function of temperature or pH. These findings establish a useful assay for screening potential cancer drugs that bind in a specific way to MGL that can be carried out more rapidly and with significantly smaller amounts of protein than other assays for the dynamic motion of the drug target.Grant Funding Source: Supported by grants DA09158, DA00493, DA03801 and DA07312