Ring current shifts in (19)F-NMR of membrane proteins

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 (19)F-NMR probes include high sensitivity of the (19)F chemical shifts to variations in the non-covalent environment. Nonetheless, in studies of G protein-coupled receptors (GPCR) we encountered situations where (19)F 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 (19)F-NMR probes used in GPCRs. Analysis of previously reported (19)F-NMR data on the β2-adrenergic receptor and mammalian rhodopsin showed that all (19)F-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 (19)F chemical shifts, there is thus an indication that their contributions are significant when studying activation processes in GPCRs, since the observed activation-related (19)F-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 (19)F-NMR studies of GPCR activation, and novel information may be obtained on the nature of conformational rearrangements near the (19)F-labels. It will then also be interesting to see if the presently indicated role of ring current shifts in membrane protein studies with (19)F-NMR markers can be substantiated by a more extensive data base resulting from future studies.