Structural Studies of Neuropeptide Y and NPY-[18-36] in the Presence of Neuronal Lipid Membrane Mimics

Abstract

Neuropeptide Y (NPY), a 36-residue long polypeptide and one of the most abundant neuropeptides found in the mammalian nervous system, activates G-protein coupled receptors. As a neurotransmitter, it helps regulate metabolism and the cardiovascular system. It has also been shown to have antimicrobial properties. To better understand multiple functions of NPY, it is important to characterize its structural features under physiologically-relevant conditions. Previous studies have shown that NPY interacts strongly with phospholipids that mimic neuronal cell membranes. In this regard, we recently used CD and solid-state NMR to show that 2:1PC:PS (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine/ 1-palmitoyl 2-oleoyl phosphatidylserine), a suitable lipid mimic for neuronal membranes, could be used for high-resolution structural studies of NPY and antagonist NPY-[18-36].In this research, we have used solid-state NMR more extensively on NPY and NPY-[18-36] to investigate their high-resolution structures and dynamics in the presence of 2:1 POPC/POPS bilayers. Two-dimensional heteronuclear correlation solid-state NMR spectra have been collected on 15N-amide backbone labeled peptides in the presence of aligned lipid bilayers. 15N chemical shifts and 15N-1H dipolar couplings have been used to obtain the topological orientation and preliminary backbone structures of the peptides. The data indicate that the alpha-helical portion of the peptides adopt an in-plane bilayer orientation and diffusive rapidly in the plane of the membrane. To investigate the membrane-binding and membrane-disrupting abilities of NPY and NPY-[18-36], we performed CD and dye leakage experiments on samples containing vesicles of various lipid compositions and different peptide concentrations. We will discuss the results in terms of the peptide's ability to disrupt lipid bilayers mimicking mammalian and microbial cells. Overall, these studies provide valuable knowledge to help elucidate the molecular basis of NPY's multiple functions in the human body.