These pdb files show the extracellular domains (ECDs) of the human calcitonin (CTR) and amylin 1 receptors (AMY1R). Additional models of the ECD of the AMY1R with either CGRP or amylin bound are also included.
CTR and CLR full length sequences were aligned using Tcoffee (Poirot et al., 2003). A multiple template design was used to generate the CTR ECD. CLR ECD, chain A from 3N7S, was used as a template structure for CTR ECD. The first loop domain of chain A (3N7S) is not elucidated. Therefore, the loop domain of 3N7S (chain B) was used as the template for this loop domain in the CTR ECD. Modeller9v8 (Sali and Blundell, 1993) was used to generate 500 models. The models were ranked by the Modeller9v8 energy objective function. The top 10 structures were retained and the stereochemical quality was assessed by PROCHECKv3.5.4 (Laskowski et al., 2001). Based on overall and residue-by-residue geometry a structure was selected.The ProPka program (Li et al., 2005) via the PDBQPR server (see Dolinsky et al., 2007) was used to assign the protonation states of the titratable groups in CTR ECD, using the CHARMM parameters set at pH 7.0. The CHARMM (c35b3) module Screened Coulomb Potentials Implicit Solvent Model (SCPISM) was used to minimise the model. 100 steps of steepest descent were conducted followed by adopted basis Newton-Raphson minimisation until convergence was met.
To model CGRP and amylin bound to the AMY receptor, a similar approach was used to that for the modelling of the receptor without ligand, but the structure of a CGRP analog bound to the CGRP receptor ECD was used as a template (PDB ID: 4RWG). This was mutated in-silico to give the structures of either human CGRP or amylin. Subsequent steps were as described above.
Funding: Maurice Wilkins Centre for Molecular Biodiscovery, Maurice and Phyllis Paykel Trust, University of Auckland doctoral scholarship, National Heart Foundation of New Zealand, National Institute of Health (R01GM104251), Wellcome Trust (091496), National Health and Medical Research Council of Australia (1061044, 1055134)