Knowledge of the high-resolution structures of GPCRs is a crucial first step in understanding their functions in cellular membranes. Rhodopsin is a prototype for the Family-A GPCRs. Despite several reported crystal structures for the rhodopsin Meta-II state, the conformation and orientation of the retinal chromophore in its binding pocket are still ambiguous [1]. Here, we investigated rhodopsin in aligned membranes using solid-state 2H NMR spectroscopy and QM/MM simulations. Rhodopsin was regenerated with retinal 2H labeled at the C5-, C9-, or C13-methyl groups, then recombined into aligned phospholipid membranes, and trapped in the light-activated state [2]. By fitting theoretical 2H NMR line-shapes to experimental spectra, we determined the orientation of the 2H-labeled methyl groups to the membrane normal, which served as restraints for structural calculations. We discovered that two different retinal structures are possible in the active Meta-II state. The first resembles the proposed active X-ray structures, with both the polyene chain and the β-ionone ring rotated about the molecular long-axis. The second structure has the same position of the β-ionone ring, whereas the polyene chain remains unflipped. In QM/MM simulations, four models were tested with different orientations of the chromophore and different protonation states of Glu113 and Glu181. We discovered two possible structures (“flipped” and “unflipped”) without significant steric clashes in the binding pocket of active rhodopsin. Perhaps most arresting, solid-state 2H NMR spectroscopy yields the sequence of activating retinal structural changes (dark, Meta-I, and Meta-II states) for the first time. An ensemble of conformational substates characterizes dynamically active GPCRs, and enables their highly efficient interactions with G-proteins and downstream effectors [3]. [1] J. Feng et al. (2015) Biophys.J. 108, 2754. [2] A.V. Struts et al. (2015) Methods Mol.Biol. 1271,133. [3] Perera et al. (2018) J.Phys.Chem.Lett. 9, 7064.