Probing the Y2 Receptor on Transmembrane, Intra- and Extra-Cellular Sites for EPR Measurements.

ABSTRACT: The function of G protein-coupled receptors is intrinsically linked to their conformational dynamics. In conjugation with site-directed spin labeling, electron paramagnetic resonance (EPR) spectroscopy provides powerful tools to study the highly dynamic conformational states of these proteins. Here, we explored positions for nitroxide spin labeling coupled to single cysteines, introduced at transmembrane, intra- and extra-cellular sites of the human neuropeptide Y2 receptor. Receptor mutants were functionally analyzed in cell culture system, expressed in Escherichia coli fermentation with yields of up to 10 mg of purified protein per liter expression medium and functionally reconstituted into a lipid bicelle environment. Successful spin labeling was confirmed by a fluorescence assay and co

SUBMITTER: Laugwitz JM 

PROVIDER: S-EPMC7571237 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature