Comparative conformational studies on cyclic hexapeptides corresponding to message sequence His-Phe-Arg-Trp of...

Solution conformation of cyclo(Gly1-His2-Phe3-Arg4-Trp5-Gly6) and its D-Phe analog corresponding to the message sequence [Gly-alpha-MSH5-10] of alpha-MSH has been studied by 1D and 2D proton magnetic resonance spectroscopy in dimethyl sulfoxide (DMSO)-d6 solution and in a DMSO-d6/H2O cryoprotective mixture. The NMR data for both the analogs in solution at 300 K cannot be interpreted based on a single ordered conformation, as evidenced by the broadening of only -NH resonances as well as the temperature coefficients of the amide protons. An analysis of the nuclear Overhauser effect (NOE) cross-peaks in conjunction with temperature coefficient data indicates an equilibrium of multiple conformers with a substantial population of particular conformational states at least in the D-analog. The molecular dynamics simulations without and with NOE constraints also reveal numerous low-energy conformers with two gamma-turns, a gamma-turn and a beta-turn, two beta-turns, etc. for both the analogs. The observed NMR spectra can be rationalized by a dynamic equilibrium of conformers characterized by a gamma-bend at Gly6, two gamma-bends at Phe3 and Gly6 and a conformer with a single beta-turn and a gamma-bend for the L-Phe analog. On the other hand, a conformation with two fused beta-turns around the two tetrads His2-D-Phe3-Arg4-Trp5 and Trp5-Gly6-Gly1-His2 dominates the equilibrium mixture for the D-Phe analog. For the D-Phe analog, the experimentally observed average conformation is corroborated by molecular dynamics simulations as well as by studies in cryoprotective solvent.