Soft matter, namely, water and lipids, comprises an essential part of the native environment of membrane proteins. However, the mechanism of how lipids and water interact with proteins remains poorly understood. To uncover these questions, we use rhodopsin which is an archetypical G-protein-coupled receptor (GPCR) that serves as a model to understand soft matter effects. Upon photoactivation, the chromophore retinal isomerizes from 11- cis to all- trans within the rhodopsin orthosteric site, triggering conformational changes into several intermediates, including preactive metarhodopsin-I and active metarhodopsin-II states. We previously demonstrated that the activation equilibrium is sensitive to dehydration of the protein interior and lipid-protein interactions as explained by the flexible surface model. 1,2 Here, we test this sponge model of rhodopsin activation using osmolytes to probe the effects of dehydration on the binding of transducin mimetic peptides to rhodopsin in various lipid environments. 1 UV-visible spectroscopy was used to measure rhodopsin activation, and a binding model was employed to calculate the dissociation constant of the peptides. We hypothesize that binding and release of the transducin G-protein is coupled to rhodopsin hydration cycling. Notably, we discovered that osmolytes affect rhodopsin activation, with small osmolytes penetrating the binding pocket and increasing activation. Furthermore, we found that osmolytes do not compete with binding of these mimetic peptides as the binding constant is unaffected by small osmolytes. We also observe that rhodopsin in unsaturated recombinant lipids has a higher binding affinity to these peptides than its native membrane, hinting that curvature forces are involved in G-protein binding. Unveiling water and lipid effects on transducin peptide binding to rhodopsin offers medical applications, from pharmaceutical drug development to deciphering disease mechanisms. 1 Chawla, U., et al. (2021) Angew. Chem. Int. Ed. 60 , 2288. 2 Fried, S. D. E., et al. (2022) PNAS 119 , e2117349119.
Luu et al. (Sun,) studied this question.