Retinal orientation and interactions in rhodopsin reveal a two-stage trigger mechanism for activation

The 11-cis retinal chromophore is tightly packed within the interior of the visual receptor rhodopsin and isomerizes to the all-trans configuration following absorption of light. The mechanism by which this isomerization event drives the outward rotation of transmembrane helix H6, a hallmark of activated G protein-coupled receptors, is not well established. To address this question, we use solid-state NMR and FTIR spectroscopy to define the orientation and interactions of the retinal chromophore in the active metarhodopsin II intermediate. Here we show that isomerization of the 11-cis retinal chromophore generates strong steric interactions between its β-ionone ring and transmembrane helices H5 and H6, while deprotonation of its protonated Schiff’s base triggers the rearrangement of the hydrogen-bonding network involving residues on H6 and within the second extracellular loop. We integrate these observations with previous structural and functional studies to propose a two-stage mechanism for rhodopsin activation

Free backbone carbonyls mediate rhodopsin activation

Conserved prolines in the transmembrane helices of G-protein-coupled receptors (GPCRs) are often considered to function as hinges that divide the helix into two segments capable of independent motion. Depending on their potential to hydrogen-bond, the free C=O groups associated with these prolines can facilitate conformational flexibility, conformational switching or stabilization of the receptor structure. To address the role of conserved prolines in family A GPCRs through solid-state NMR spectroscopy, we focus on bovine rhodopsin, a GPCR in the visual receptor subfamily. The free backbone C=O groups on helices H5 and H7 stabilize the inactive rhodopsin structure through hydrogen-bonds to residues on adjacent helices. In response to light-induced isomerization of the retinal chromophore, hydrogen-bonding interactions involving these C=O groups are released, thus facilitating repacking of H5 and H7 onto the transmembrane core of the receptor. These results provide insights into the multiple structural and functional roles of prolines in membrane proteins

Amino acid conservation and interactions in rhodopsin: Probing receptor activation by NMR spectroscopy

Rhodopsin is a classical two-state G protein-coupled receptor (GPCR). In the dark, its 11-cis retinal chromophore serves as an inverse agonist to lock the receptor in an inactive state. Retinal-protein and protein-protein interactions have evolved to reduce the basal activity of the receptor in order to achieve low dark noise in the visual system. In contrast, absorption of light triggers rapid isomerization of the retinal, which drives the conversion of the receptor to a fully active conformation. Several specific protein-protein interactions have evolved that maintain the lifetime of the active state in order to increase the sensitivity of this receptor for dim-light vision in vertebrates. In this article, we review the molecular interactions that stabilize rhodopsin in the dark-state and describe the use of solid-state NMR spectroscopy for probing the structural changes that occur upon light-activation. Amino acid conservation provides a guide for those interactions that are common in the class A GPCRs as well as those that are unique to the visual system. This article is part of a Special Issue entitled: Retinal Proteins - You can teach an old dog new tricks. © 2013 Elsevier B.V