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A Conserved Proline Hinge Mediates Helix Dynamics and Activation of Rhodopsin.

ABSTRACT: Despite high-resolution crystal structures of both inactive and active G protein-coupled receptors (GPCRs), it is still not known how ligands trigger the large structural change on the intracellular side of the receptor since the conformational changes that occur within the extracellular ligand-binding region upon activation are subtle. Here, we use solid-state NMR and Fourier transform infrared spectroscopy on rhodopsin to show that Trp265^6.48 within the CWxP motif on transmembrane helix H6 constrains a proline hinge in the inactive state, suggesting that activation results in unraveling of the H6 backbone within this motif, a local change in dynamics that allows helix H6 to swing outward. Notably, Tyr301^7.48 within activation switch 2 appears to mimic the negative allosteric sodium ion found in other family A GPCRs, a finding that is broadly relevant to the mechanism of receptor activation.

SUBMITTER: Pope AL

PROVIDER: S-EPMC7484155 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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A Conserved Proline Hinge Mediates Helix Dynamics and Activation of Rhodopsin.

Pope Andreyah L AL Sanchez-Reyes Omar B OB South Kieron K Zaitseva Ekaterina E Ziliox Martine M Vogel Reiner R Reeves Philip J PJ Smith Steven O SO

Structure (London, England : 1993) 20200528 9

Despite high-resolution crystal structures of both inactive and active G protein-coupled receptors (GPCRs), it is still not known how ligands trigger the large structural change on the intracellular side of the receptor since the conformational changes that occur within the extracellular ligand-binding region upon activation are subtle. Here, we use solid-state NMR and Fourier transform infrared spectroscopy on rhodopsin to show that Trp265<sup>6.48</sup> within the CWxP motif on transmembrane h ...[more]

PMID: 32470317

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Project description:Rhodopsin is a canonical member of class A of the G protein-coupled receptors (GPCRs) that are implicated in many of the drug interventions in humans and are of great pharmaceutical interest. The molecular mechanism of rhodopsin activation remains unknown as atomistic structural information for the active metarhodopsin II state is currently lacking. Solid-state (2)H NMR constitutes a powerful approach to study atomic-level dynamics of membrane proteins. In the present application, we describe how information is obtained about interactions of the retinal cofactor with rhodopsin that change with light activation of the photoreceptor. The retinal methyl groups play an important role in rhodopsin function by directing conformational changes upon transition into the active state. Site-specific (2)H labels have been introduced into the methyl groups of retinal and solid-state (2)H NMR methods applied to obtain order parameters and correlation times that quantify the mobility of the cofactor in the inactive dark state, as well as the cryotrapped metarhodopsin I and metarhodopsin II states. Analysis of the angular-dependent (2)H NMR line shapes for selectively deuterated methyl groups of rhodopsin in aligned membranes enables determination of the average ligand conformation within the binding pocket. The relaxation data suggest that the beta-ionone ring is not expelled from its hydrophobic pocket in the transition from the pre-activated metarhodopsin I to the active metarhodopsin II state. Rather, the major structural changes of the retinal cofactor occur already at the metarhodopsin I state in the activation process. The metarhodopsin I to metarhodopsin II transition involves mainly conformational changes of the protein within the membrane lipid bilayer rather than the ligand. The dynamics of the retinylidene methyl groups upon isomerization are explained by an activation mechanism involving cooperative rearrangements of extracellular loop E2 together with transmembrane helices H5 and H6. These activating movements are triggered by steric clashes of the isomerized all-trans retinal with the beta4 strand of the E2 loop and the side chains of Glu(122) and Trp(265) within the binding pocket. The solid-state (2)H NMR data are discussed with regard to the pathway of the energy flow in the receptor activation mechanism.

Dataset Information

A Conserved Proline Hinge Mediates Helix Dynamics and Activation of Rhodopsin.

Publications

A Conserved Proline Hinge Mediates Helix Dynamics and Activation of Rhodopsin.

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