Ring current shifts in ¹⁹F-NMR of membrane proteins

Abstract

Fluorine-19 NMR markers are attractive reporter groups for use in studies of complex biomacromolecular systems, in particular also for studies of function-related conformational equilibria and rate processes in membrane proteins. Advantages of ¹⁹F-NMR probes include high sensitivity of the ¹⁹F chemical shifts to variations in the non-covalent environment. Nonetheless, in studies of G protein-coupled receptors (GPCR) we encountered situations where ¹⁹F chemical shifts were not responsive to conformational changes that had been implicated by other methods. This prompted us to examine possible effects of aromatic ring current fields on the chemical shifts of ¹⁹F-NMR probes used in GPCRs. Analysis of previously reported ¹⁹F-NMR data on the β₂-adrenergic receptor and mammalian rhodopsin showed that all ¹⁹F-labeling sites which manifested conformational changes are located near aromatic residues. Although ring current effects are small when compared to other known non-covalent effects on ¹⁹F chemical shifts, there is thus an indication that their contributions are significant when studying activation processes in GPCRs, since the observed activation-related ¹⁹F-NMR chemical shifts are comparable in size to the calculated ring current shifts. Considering the impact of ring current shifts may thus be helpful in identifying promising indigenous or engineered labeling sites for future ¹⁹F-NMR studies of GPCR activation, and novel information may be obtained on the nature of conformational rearrangements near the ¹⁹F-labels. It will then also be interesting to see if the presently indicated role of ring current shifts in membrane protein studies with ¹⁹F-NMR markers can be substantiated by a more extensive data base resulting from future studies.