Abstract
Calcitonin is a peptide hormone consisting of 32 amino acid residues and the calcitonin receptor is a Class B G protein-coupled receptor (GPCR). The crystal structure of the human calcitonin receptor ectodomain (CTR ECD) in complex with a truncated analogue of salmon calcitonin ([BrPhe(22)]sCT(8-32)) has been determined to 2.1-Å resolution. Parallel analysis of a series of peptide ligands showed that the rank order of binding of the CTR ECD is identical to the rank order of binding of the full-length CTR, confirming the structural integrity and relevance of the isolated CTR ECD. The structure of the CTR ECD is similar to other Class B GPCRs and the ligand binding site is similar to the binding site of the homologous receptors for the calcitonin gene-related peptide (CGRP) and adrenomedulin (AM) recently published (Booe, J. M., Walker, C. S., Barwell, J., Kuteyi, G., Simms, J., Jamaluddin, M. A., Warner, M. L., Bill, R. M., Harris, P. W., Brimble, M. A., Poyner, D. R., Hay, D. L., and Pioszak, A. A. (2015) Mol. Cell 58, 1040-1052). Interestingly the receptor-bound structure of the ligand [BrPhe(22)]sCT(8-32) differs from the receptor-bound structure of the homologous ligands CGRP and AM. They all adopt an extended conformation followed by a C-terminal β turn, however, [BrPhe(22)]sCT(8-32) adopts a type II turn (Gly(28)-Thr(31)), whereas CGRP and AM adopt type I turns. Our results suggest that a type II turn is the preferred conformation of calcitonin, whereas a type I turn is the preferred conformation of peptides that require RAMPs; CGRP, AM, and amylin. In addition the structure provides a detailed molecular explanation and hypothesis regarding ligand binding properties of CTR and the amylin receptors.
Highlights
Calcitonin (CT)3 is a peptide hormone that possesses the ability to lower the calcium plasma concentration [1]
The structures of several Class B G protein-coupled receptor (GPCR) ECDs in complex with and without ligands have been determined including the ECDs of corticotropin-releasing factor receptor [5], pituitary adenylate cyclase-activating polypeptide receptor [6], glucose-dependent insulinotropic polypeptide receptor [7], parathyroid hormone-1 receptor [8], glucagon-like peptide-1 receptor [9], calcitonin receptor-like receptor (CLR) [10], and the glucagon receptor [11]
The CLR ECD was used as the model for molecular replacement (CLR ECD PDB entry 3N7S, chain A only), and it shares ϳ60% amino acid residue sequence identity with the crystallized calcitonin receptor ectodomain (CTR ECD) of this study
Summary
The molecular details that govern ligand binding and specificity of CTR have not previously been clarified by high resolution structure determination. Likewise the change of ligand specificity of CTR enforced by the interaction with RAMP is not fully understood. In this study we determined the crystal structure of the CTR ECD in complex with the peptide ligand [BrPhe22]sCT8 –32 ([BrPhe22] ϭ 4-bromo-L-phenylalanine at position 22). The molecular details of the interaction between sCT and the CTR ECD is revealed by the structure and provides new information regarding the structure of sCT in the receptor-bound state and the molecular determinants of ligand specificity of this Class B receptor subclass, in particular the receptors for calcitonin and amylin
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