The NMR-Derived Conformation of Orexin-A: An Orphan G-Protein Coupled Receptor Agonist Involved in Appetite Regulation and Sleep

Abstract

The conformation of orexin-A, an orphan G-protein coupled receptor agonist (the human sequence is: has been determined when bound to sodium dodecylsulphate-d25 (SDS) micelles by 1H and 13C NMR and molecular modeling. Orexin-A has been implicated in sleep-wakefulness and feeding regulation. The conformational preference of orexin-A consists of a short helical section, involving Asp5 to Gln9 that makes up helix I, followed by a bend from Lys10 to Ser13. Residues Leu16 to Gly22 make up helix II. The conformation of orexin-A can now be used to explain the results of earlier Ala substitution mutagenesis experiments (J. G. Darker, et al. Bioorg. Med. Chem. Lett. 11, 737–740 (2001); S. Ammoun, et al. J. Pharmacol. Expt. Ther. 305, 507–514 (2003)). Darker et al., working with orexin-A (15–33) amide, observed a significant drop in functional potency at the OX1R receptor when Leu16, Leu19, Leu20, His26, Gly29, Ile30, Leu31, Thr32, and Leu33 were replaced by Ala. Ammoun et al. identified three areas of interest, which were the same for OX1R and OX2 R receptors, as amino acids 15–17, 20 and 25–26 with the most marked reduction in activity being produced by the replacement of Leu20 by Ala. We suggest that Leu16, Leu19, and Leu20, which are in helix II, are likely responsible for binding orexin-A to the surface of the micelle.