Conformational Characterization of Tachykinin Neuropeptides: Role of the Polyproline II Structure

Abstract

Substance P (SP), Neurokinin A (NKA) and Scyliorhinin I (Scyl) peptides belong to the Tachykinin family and are agonists for Neurokinin1 (NK1) GPCR receptor with different potent activities. These Tachykinins are involved in several physiological processes and some neurodegenerative disorders, what make them relevant therapeutically agents. The characterization of the active peptide conformations of the Tachykinins is essential for the elucidation of the mechanism of action and for the design of drugs. To address the molecular basis for the peptide recognition by the NK1 receptor we studied the conformation of SP, NKA and Scyl in solution and membrane-mimic environments: micelles and liposomes, by using CD and FTIR spectroscopies. The analysis of CD data revealed that the three peptides form a partially α helical structure in the presence of the negatively charged micelles and vesicles, but not in zwitterionic DMPC. By performing CD spectra at increasing temperatures we demonstrated that in an aqueous environment SP, NKA and Scyl form extended polyproline II (PPII) helical structure. The same structure was found in the membrane mimics, which do not induce formation of α helical conformation of the peptides. FTIR experiments performed in D2O support the presence of PPII conformation. Further we questioned whether the formation of PPII needs peptide binding to the membrane or it is a peptide intrinsic property, by using fluorescence techniques. We propose that the peptide-membrane interaction is a two-step process involving first electrostatic interactions through PPII structure, followed by the folding and insertion of the alpha-helical segment.