Developing a DeSipher method for conformational change detection of membrane receptor complex at residue resolution

Characterization of the dynamic conformational changes in membrane protein signaling complexes by nuclear magnetic resonance (NMR) spectroscopy remains challenging. Here, we developed "DeSipher" technology to site-specifically incorporate 4-trimethylsilyl phenylalanine (TMSiPhe) into proteins and measured the conformational changes in the signaling complex by 1H NMR. Notably, crystallographic analysis revealed structural changes that reshaped the TMSiPhe-specific amino-acyl tRNA synthetase to accommodate the large trimethylsilyl (TMSi) group. The unique 1H NMR chemical shift of TMSiPhe and highly efficient incorporation of TMSiPhe enabled facile assignment of the associated NMR signal. We further showed that extracellular ligands induced conformational changes located in the polar core or ERK interaction site of beta-arrestin-1 via direct receptor transmembrane core interactions in the phospho-beta2AR/beta-arrestin-1 signaling complex. These observations provided direct delineation and key mechanism insights that multiple receptor ligands were able to induce distinct arrestin functionally relevant conformational changes at residue levels.