Solid-State ²H NMR Structure of Retinal in Metarhodopsin I

Abstract

The structural and photochemical changes in rhodopsin due to absorption of light are crucial for understanding the process of visual signaling. We investigated the structure of trans-retinal in the metarhodopsin I photointermediate (MI), where the retinylidene cofactor functions as an antagonist. Rhodopsin was regenerated using retinal that was (2)H-labeled at the C5, C9, or C13 methyl groups and was reconstituted with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine. Membranes were aligned by isopotential centrifugation, and rhodopsin in the supported bilayers was then bleached and cryotrapped in the MI state. Solid-state (2)H NMR spectra of oriented rhodopsin in the low-temperature lipid gel state were analyzed in terms of a static uniaxial distribution (Nevzorov, A. A.; Moltke, S.; Heyn, M. P.; Brown, M. F. J. Am. Chem. Soc. 1999, 121, 7636-7643). The line shape analysis allowed us to obtain the methyl bond orientations relative to the membrane normal in the presence of substantial alignment disorder (mosaic spread). Relative orientations of the methyl groups were used to calculate effective torsional angles between the three different planes that represent the polyene chain and the beta-ionone ring of retinal. Assuming a three-plane model, a less distorted structure was found for retinal in MI compared to the dark state. Our results are pertinent to how photonic energy is channeled within the protein to allow the strained retinal conformation to relax, thereby forming the activated state of the receptor.

Solid-state 2 H NMR relaxation illuminates functional dynamics of retinal cofactor in membrane activation of rhodopsin

“…Despite that three methyl environments are found for the retinal ligand in the dark state, upon 11-cis to trans isomerization ≈ two methyl sites are evident. Such a loss of retinal strain is consistent with two major planes of unsaturation following light absorption (1,34,(38)(39)(40). Application of transition state theory then allows us to connect the T 1Z results with the energetic parameters for retinal bound to rhodopsin (Table 1).…”

250GHz CW gyrotron oscillator for dynamic nuclear polarization in biological solid state NMR

“…Two research areas in which high resolution MAS experiments have proved especially successful are studies of amyloid fibrils [37,[51][52][53][54][55][56][57] and membrane proteins [42,[58][59][60][61][62][63][64][65][66][67][68][69][70][71][72][73][74][75]. However, in both of these cases, low sensitivity currently limits the information that can be gleaned from the spectra.…”

Two protonation switches control rhodopsin activation in membranes

“…Within milliseconds several inactive intermediates are formed, such as Batho, BSI, Lumi, and Meta I, that can be examined by using time-resolved (3) or cryotrapping techniques (4). The early transitions involve mainly a relaxation of the isomerized retinal chromophore (5,6), with only minor changes to the ␣-helix bundle of the receptor protein as revealed by electron crystallography of the Meta I state (7). Only in the transition from Meta I to the active receptor conformation Meta II is a rearrangement of the helix bundle observed, involving tilt movements of H6 (8)(9)(10) and presumably also of H5 (11).…”