Patent ID: 9069700
Filing Date: 2015-06-30
Classification: C07K,G01N,G16B

Abstract:
1. A method of obtaining structures for active, inactive, and intermediate active forms of rhodopsin for identifying or designing ligands that bind to the intermediate rhodopsin forms comprising: (a) obtaining three-dimensional structural coordinates for the crystal structure of rhodopsin in the ground state; (b) pivoting transmembrane helix 3 (TM3) about the Cα carbon of Cys110 such that Cys140 is at a set distance from transmembrane 6 (TM6), while keeping the distance of TM3 to transmembrane helix 2 (TM2) at 5 Å or more, wherein the set distance from TM6 is selected from the group consisting of: 4.3 Å to form activated metarhodopsin I intermediate of rhodopsin, 6.8 Å to form activated metarhodopsin Ib intermediate of rhodopsin, and 9.0 Å to form activated metarhodopsin I (c) pivoting transmembrane helix 4 (TM4) about Gly174 such that Glu150 is at a set distance from transmembrane helix 5 (TM5), wherein the set distance from TM5 is selected from the group consisting of: 7.4 Å to form activated metarhodopsin I intermediate of rhodopsin, 12.1 Å to form activated metarhodopsin Ib intermediate of rhodopsin, and 17.1 Å to form activated metarhodopsin I (d) optimizing the structural model of step (c) at 300K by molecular dynamics and energy minimization such that the Cα carbons of the amino acids are fixed, wherein amino acid residues have the same numbering as in bovine rhodopsin, and wherein transmembrane helixes 1, 2, and 7 are not subjected to a conformational change; (e) optionally further comprising, to form activated metarhodopsin II intermediate of rhodopsin from metarhodopsin I, metarhodopsin Ib, or metarhodopsin I (f) binding a previously identified or designed ligand to the activated metarhodopsin I, metarhodopsin Ib, metarhodopsin I