Solution Conformations of Potent Bicyclic Antagonists of Oxytocin by Nuclear Magnetic Resonance Spectroscopy and Molecular Dynamics Simulations

Abstract

The solution conformations of two potent bicyclic antagonists [dPen1, cyclo(Glu4,Lys8)]OT were studied by a combined use of 1H and 13C NMR spectroscopy in DMSO and molecular dynamics (MD) simulations. NMR data have suggested a model for the three-dimensional (3D) structure of the bicyclic analogues of OT (OT-BC) with a β-turn at the Tyr2 and Ile3 residues, and with a cis amide bond between Cys6 and Pro7. A 3D structure containing a type III β-turn at Tyr2-Ile3 has been shown to be consistent with NMR data. This structure was proposed as a model of the solution conformation of OT-BC and extensively tested by MD simulations with the AMBER force field. MD simulations at 300 K with NMR derived distance and φ torsion angle constraints demonstrated the consistency of this model with NMR data, and its stability was further demonstrated by non-constrained MD simulations. Dynamic properties of the 3D structure were explored by high-temperature MD at 500 K. Conformational transitions induced by a constrained rotation around the S−S bond revealed relatively low potential energy barriers (30 to 50 kJ/mol) between equilibrium left-handed and right-handed conformers of the disulfide bridge in OT-BC. A dynamic model of the solution structure of OT-BC with the relatively stable backbone conformation and a fast conformational equilibrium in the disulfide bridge and lactam bridge moieties was proposed as a result of the extensive MD simulations. The solution structure of OT-BC is consistent with structure−activity relations of peptide and non-peptide antagonists of OT. In particular, a β-turn at Tyr2-Ile3 seems to be the common feature responsible for antagonist interaction with the uterine receptor of OT. On the other hand, the 3D structure of OT-BC differs considerably from the crystal and solution structures of OT analogues with agonist activity. Therefore, this study supports the hypothesis of different modes of receptor binding for agonists and antagonists of OT. The model of 3D structure of OT-BC proposed in this study may be used as a template for the rational design of peptide and non-peptide antagonists of oxytocin.

Examination of structural characteristics of the potent oxytocin antagonists [dPen1,Pen6]-OT and [dPen1,Pen6, 5-tBuPro7]-OT by NMR, raman, CD spectroscopy and molecular modeling

“…Due to its small population, most of the hydrogen amide resonances for the minor cis-isomer conformation of 1 were not detected. Temperature coefficients consistent with solvent shielded hydrogens were previously observed in the studies of antagonist analog [Pen 1 ]-OT (pA 2 = 6.9) for Ile 3 and Asn 5 [17]; and in the bicyclic antagonists [Mpa 1 ,cyclo(Glu 4 ,Lys 8 )]-OT (pA 2 = 8.2) and [dPen 1 ,cyclo(Glu 4 ,Lys 8 )]-OT (pA 2 = 8.7) for Ile 3 and Glu 4 [23]. Temperature coefficients were also calculated for the NH 2 amide proton resonances of Gln 4 , Asn 5 and Gly 9 in analogs 1 and 2 (data not shown) and the δ/ T values ranged from −6.5 to −4.3 × 10 −3 ppm/K indicating the absence of strong hydrogen bonding.…”

“…the major (trans-) conformers of 1 and 2 suggested along with the Raman and CD data that a subset of lower-energy conformations existed with a specific disulfide geometry. Computational methods have been used extensively to search for preferred conformations for oxytocin analogs, and have resulted in the discovery of several families of stable conformations [23,[79][80][81][82][83]. In a similar fashion, a conformational search was undertaken to assess whether or not the observed data corresponded to a unique set of conformations for both analogs.…”

“…possessed the amide N -terminal to proline 7 in the transisomer conformation, an NMR spectroscopic study of OT (3) demonstrated that the amide N -terminal to Pro 7 could adopt up to 10% cis-isomer in solution [21,22]. Furthermore, conformational analyses of the potent bicyclic OT antagonists [Mpa 1 ,cyclo(Glu 4 ,Lys 8 )]-OT and [dPen 1 ,cyclo(Glu 4 ,Lys 8 )]-OT by NMR spectroscopy (DMSO) and molecular modeling revealed the disulfide bond to be flexible and the prolyl amide locked in the cis-isomer conformation [23]. Comparison of these bicyclic analogs and other OT analogs has led to the hypothesis that the prolyl amide trans-isomer is necessary for agonist activity and that the cis-isomer may favor antagonism [24].…”

Conformational Sampling and Energetics of Drug-Like Molecules

“…in the context of pharmacophore studies [219] or in conjunction with NMR data [138,[220][221][222]. Distance geometry is another method of choice for these situations.…”

Preterm Labour: An Overview of Current and Emerging Therapeutics

“…corresponding acyclic compounds [250,251]. NMRspectroscopic examination of the solution structures of two bicyclic analogues in comparison with OT have shed more light on our understanding of the different modes of receptor binding for OT-R agonists and antagonists [252].…”