An Activating Helix Switch at the Rhodopsin-Transducin Interface

Peter W. Hildebrand, Alexander Rose, Klaus P. Hofmann
2012 Biophysical Journal  
a multidisciplinary approach combining computational and experiment work that leverages recent structural data, we present a model for the supramolecular structure of the rod cell disc membrane phototransduction machinery. We first show a comparison of multiple rhodopsin dimer interfaces in a model membrane using a coarse-grained molecular dynamics simulation approach accumulating more than a millisecond of simulation time. To characterize the preferred binding interface of a pair of
more » ... we determined the potentials of mean force as a function of the distance between two membraneembedded receptors. The interfaces probed include helix 4 (H4), H4/H5, H5, H6 and H1/H8. The results show that the most stable rhodopsin dimer exists in a tail-to-tail conformation, with the interface comprising transmembrane H1 and H2 at the extracellular side and amphipathic H8 at the cytoplasmic one. The existence of the H1/H8 dimer was unambiguously corroborated by crosslinking experiments in which we identified CYS316 in H8 as the site of a chemical crosslink between two rhodopsins in native ROS disc membrane using proteolysis, CNBr cleavage, and high-resolution liquid-chromatography mass-spectroscopy (LC/MS). We then show how secondary interaction surfaces appear to stabilize extended "lubricated" rows of these dimers as seen earlier in atomic force microscopy studies. The synthesis of the new rhodopsin dimer orientation with the structures of the R*/Gt complex in two different orientations and 3-D densities of the complex obtained earlier by electronmicroscopy argues for novel alternatives for the supramolecular organization of ROS membrane. The model we propose suggests the possibility for an efficient one-dimensional mechanism for Gt to search for active receptor (R*) even under low light conditions. The recent crystal structure 1 of the b 2 adrenergic receptor-Gs protein complex (b 2 AR-Gs) marks an important milestone in a two decades long race to decipher one of the most important and widely utilized signal-transduction mechanisms in biology. However, it is unclear how the relatively drastic and unexpected conformational changes evolve from the initial encounter complex. The number of compositional and conformational microstates that evolve along the reaction pathway from the initial encounter of agonist-activated receptor with the GDP-bound heterotrimeric G-protein to the nucleotide-free agonist-receptor-G protein ternary complex is potentially very large. 2 Identification of these pathways is out of reach for crystallographic experiments, and we follow a complementary approach with large-scale molecular dynamics (MD) simulations of membrane-embedded, fully solvated metarhodopsin II alone and in complex with its cognate G-protein, transducin, in GDP-bound form. Here, we compare and contrast the relaxation of two different encounter complexes, one with an orientation similar to b 2 AR-Gs, and the other 120 rotated. Interestingly, the two different pre-aligned encounter complexes yield induced-fit complexes that share a signature set of conformational changes involving highly conserved residues in the three allosterically coupled domains, the agonist-binding, G-protein-binding, and nucleotide-binding sites. How are these canonical and non-canonical binding modes possible for such a high fidelity signal-transduction system? Is it possible that Nature utilizes multiple, energetically similar states to relay the information encoded in the relatively small free energy of agonist binding across about ten nanometers distance to the nucleotide-binding pocket? It is tempting to speculate about the role of these energetically near-equivalent microstates when discussing the dominating entropic contribution to the binding free energy of agonists at the b adrenergic receptors.
doi:10.1016/j.bpj.2011.11.2825 fatcat:34epabeedrhlrb7v62viu4tqma