Conformational Analysis Of Endomorphins Analogs Modified With Side-chain, C-terminus And Backbone By NMR Spectroscopy And Molecular Modelling

Naturally occurring opioid peptides consist of two distinct biologically important parts, a N-terminal as the message sequence, and the remaining C-terminal fragment, the address sequence. Structure-activity studies showed that the message sequence needed strict requirements for a good interaction with opioid receptors. Important features both for the message and the address sequence are the presence of cationic amino group and a phenolic group in position 1, a spacer in position 2, lipophilic and aromatic residues in positions 3 and 4, and amidation at the C-terminus, which seems to be determinant for peptide stability.Endomorphin-1 (Tyr-Pro-Trp-Phe-NH₂, EM-1) and endomorphin-2 (Tyr-Pro-Phe-Phe-NH₂, EM-2) are high potent and selective endogenous ligands toμ-opioid receptor. Although they are the most small model peptides to reveal the function of opioid ligands, several data indicates that the immunomodulatory, cardiovascular, respiratory, and analgesic effects of EMs agonists can be dissociated. To develop potent analgesics with less side effects related to it, many group have reported different methods to develop the more ideal analogs of EMs.NMR in combination with molecular modelling is the main method in investigating the conformation of peptides and small proteins. The conformations of EMs and their analogs were generated by molecular modelling and dynamics with NOE restraints after the full assignment of chemical shift in peptides. In this study, we designed some analogs of EMs with modification of side chain, C-terminus and backbone, and performed ¹H and 2D NMR spectroscopy in DMSO-d6. Based on the simulation modelling with NOE restraint, we obtain the conformations of EMs analogs and docking withμ-opioid receptor to predict the complex model of ligand-receptor.We investigated a series of conformations of endomorphin-2 (EM-2) analogs substituted by naphthyl-alanine (Nal), phenylglycine (Phg) and homophenylalanine (Hfe) in the position 3 or 4. The results show that the conformations in trans isomer have varied from extended to folded as bioactivity decreases. It is suggested that the flexibility of aromatic side chain affects the backbone of EM-2 to adopt folded structures, which may block the ligands in binding toμ-opioid receptor. And a series of analogs of EMs by modified C-terminus with Bzl, OR, -ol were designed to generate conformational constrains of the fourth aromatic ring by amide bond and torsion angles. And a series of symmetry dimeric opioid peptides composed of two tetrapeptides connected 'tail to tail' by a hydrazide bridge show that innermolecular H-bond to modulate balanced agonists to develope the structure-activity relationship between the selectivity and the distance of the two tyrosine pharmacophores. Moreover, we introduced N-O turns into EM-2 to investigate the N-O turns in bioactive peptide with potent analgesic activity by turn structure existed. These studies of conformation of analogs of EMs provide a new approach to develop potent analgesics related to EMs and are a clue to find out the characteristics selectivity of different opioid receptors.