Rhodopsin-transducin heteropentamer: Three-dimensional structure and biochemical characterization

The process of vision is initiated when the G protein-coupled receptor, rhodopsin (Rho), absorbs a photon and transitions to its activated Rho* form. Rho* binds the heterotrimeric G protein, transducin (G(t)) inducing GDP to GTP exchange and G(t) dissociation. Using nucleotide depletion and affinity chromatography, we trapped and purified the resulting nucleotide-free Rho*.G(t) complex. Quantitative SDS-PAGE suggested a 2:1 molar ratio of Rho* to G(t) in the complex and its mass determined by scanning transmission electron microscopy was 221 +/- 12 kDa. A 21.6 angstrom structure was calculated from projections of negatively stained Rho*.G(t) complexes. The molecular envelope thus determined accommodated two Rho molecules together with one G(t) heterotrimer, corroborating the heteropentameric structure of the Rho*.G(t) complex. (C) 2011 Elsevier Inc. All rights reserved.