Validating the Retinal Flip of Rhodopsin using Molecular Dynamics

Abstract

Rhodopsin, the mammalian dim light photoreceptor, is the most well-characterized structural model of a G protein-coupled receptor (GPCR). Photoisomerization of its covalently bound chromophore, retinal, triggers rhodopsin activation. Spectroscopic studies of rhodopsin in the dark [1] and Meta I [2] states have definitively shown that the C9- and C13-methyl groups of retinal are oriented towards the extracellular side of the protein. However, the structures of the active Meta II state [3] and a constitutively active triple mutant [4] had a 180° rotation along the long-axis of the retinal polyene chain, leading to an orientation of the C9- and C13-methyl groups towards the cytoplasmic side of the protein. The biophysical significance of this potential flip and its role in the structural transition during activation is still unknown. We employed molecular dynamics simulations to determine the role of the retinal flip in rhodopsin activation. Rhodopsin was modeled starting with the Meta II crystal structure but in the Meta I protonation state, to favor a deactivation transition. Surprisingly, two of our four simulations produce a reverse flip of the polyene chain on the microsecond timescale. This flip is accompanied by the rotation of the Trp265 side chain, which is implicated in a “transmission switch” common to GPCR activation. A decrease of water within the retinal binding pocket is also observed, along with distinct protein hydration features concurrent with the flipping of retinal. These results provide a bridge between spectroscopic and crystallographic studies, showing that it is possible for a retinal flip to occur from Meta I to Meta II state. [1] Salgado (2004) Biochemistry 43:12819; [2] Salgado (2006) JACS 128:11067; [3] Choe (2011) Nature 471:651; [4] Deupi (2012) PNAS 109:119.