Nuclear magnetic resonance (NMR) spectroscopy is a powerful tool for investigating various dynamic features of G protein-coupled receptor (GPCR) signaling. In this Perspective, we focus on NMR techniques to characterize ligand-dependent conformational dynamics of GPCRs as well as the interaction of GPCRs with their environment and ligands. We also describe circumstances under which each technique should be applied, their advantages and disadvantages, and how they can be combined with other strategies to deepen the understanding of GPCR signaling at the molecular level.

Phosphonic acids bind strongly to quantum dots (QDs), improving stability. However, it is difficult to introduce desired ligands to QDs passivated with phosphonic acids via ligand exchange. Here, we have investigated the efficient addition of X-type ligands to wurtzite CdSe QDs passivated with phosphonic acids using Z-type ligands. Oleic acid (OA) and Cd­(OA)2 were used as examples of the desired X-type and Z-type ligands, respectively, and octadecylphosphonic acid (ODPA) was used as a passivating ligand for CdSe QDs. OA ligands were not introduced at all into the QDs when OA itself was used as the X-type ligand. However, after Z-type ligand addition, the added OA ligand accounted for 35% of the total ligands and only 20% of ODPA was dissociated from QDs. In addition, photoluminescence (PL) and its lifetime increased 6.1 and 2.8 times, respectively, indicating that Z-type ligands bound to trap sites to increase PL. Thus, we suggest that a sufficient amount of the desired ligand can be introduced, enhancing the quantum yields of CdSe QDs substantially. Our approach can be applied to various applications requiring energy input/output control and exciton lifetime extension in QDs.

Probing the protein surface accessibility of different residues is a powerful way of characterizing the overall conformation of intrinsically disordered proteins (IDPs). We present a two-dimensional (2D) time-resolved photo-CIDNP (TR-CIDNP) experiment suitable for IDP analysis. Pulse stretching of high-power laser pulses, band-selective decoupling of 13 C α , and simultaneous application of radiofrequency and laser pulses were implemented to quantitatively analyze the IDP surface at ultrahigh resolution. Comparative analysis with other methods that measure protein surface accessibility validated the newly developed method and emphasized the importance of dye charge in photo-CIDNP. Using the neutral riboflavin dye, surface accessibilities were measured to be nearly identical for the four Tyr residues of α-synuclein (α-Syn), whose 1 H α − 13 C α correlations were well-resolved in the 2D TR-CIDNP spectrum. Having confirmed the similarity between the time-resolved and steady-state photo-CIDNP results for α-Syn, we used the more sensitive latter method to show that divalent cations induce compaction of the C-terminal region and release of the Nterminal region of α-Syn. The photo-CIDNP method presented herein can be used as an orthogonal and independent method for investigating important biological processes associated with changes in the overall IDP conformation.