Discerning conformational dynamics and binding kinetics of GPCRs by 19F NMR

Abstract

¹⁹F NMR provides a way of monitoring conformational dynamics of G-protein coupled receptors (GPCRs) from the perspective of an ensemble. While X-ray crystallography provides exquisitely resolved high-resolution structures of specific states, it generally does not recapitulate the true ensemble of functional states. Fluorine (¹⁹F) NMR provides a highly sensitive spectroscopic window into the conformational ensemble, generally permitting the direct quantification of resolvable states. Moreover, straightforward T₁- and T₂-based relaxation experiments allow for the study of fluctuations within a given state and exchange between states, on timescales spanning nanoseconds to seconds. Conveniently, most biological systems are free of fluorine. Thus, via fluorinated amino acid analogues or thiol-reactive fluorinated tags, F or CF₃ reporters can be site specifically incorporated into proteins of interest. In this review, fluorine labeling protocols and ¹⁹F NMR experiments will be presented, from the perspective of small molecule NMR (i.e. drug or small molecule interactions with receptors) or macromolecular NMR (i.e. conformational dynamics of receptors and receptor–G-protein complexes).