Neuropeptide Y

Abstract

The 36-amino-acid neuropeptide Y (human), which is one of the most potent vasoconstrictors and which exhibits a number of other biological functions, has been synthesized using automated peptide synthesis. The optimized method, using 9-fluorenylmethoxycarbonyl protecting and singlestep coupling, yielded the crude product in 90% purity allowing for single-step reversed-phase HPLC purification to >98% purity and a high overall yield (50%). The hormone was characterized by several chromatographic methods, ion-spray mass spectroscopy and Edman degradation. The conformation of human neuropeptide Y was examined by CD, NMR and computer simulations. The CD measurements in trifluoroethanol/water (9: 1) show a large percentage of a-helix. Variation of concentration, from 0.5 pM increasing up to the 1 mM used for NMR measurements, indicates no evidence for aggregation. In the same solvent system, the NMR line widths were very broad and therefore the resonance assignment was achieved with the exclusive use of two-dimensional NOE spectra. The 248 clearly distinguishable NOES from the NMR study were used in distance geometry calculations and the resulting structures were refined with restrained molecular dynamics. The results indicate an a-helix extending from Argl9 to Gln34. For the N-terminal half of the molecule no regular structure was observed.Neuropeptide Y is a 36-amino-acid C-terminally amidated peptide isolated from porcine brain by Tatemoto et al. in 1982 [l]. It is widely distributed in the central and peripheral nervous system [2, 31. Within the central nervous system neuropeptide Y is involved in appetite [4], stress [5] and blood pressure [6]. In the peripheral nervous system neuropeptide Y has been shown to be a potent and long-lasting vasoconstrictor [7], acting as a cotransmitter and regulator at noradrenergic neurons [8]. Recent studies have indicated that neuropeptide Y has an analgesic effect [9].The primary structure of porcine neuropeptide Y has been determined by sequencing [lo] and synthesis [ll]. Human neuropeptide Y has a Met residue at position 17 instead of Leu. Earlier solid-phase syntheses of porcine neuropeptide Y were performed using Na-tert-butyloxycarbonyl (Boc) Abbreviutions. ADPV, 5-(4'-aminomethyl-3',5'-dimethoxyphenoxy)valeric acid; BOP, benzotriazol-1-yl-oxy-tris(dimethy1amino)-phosphonium hexafluorophosphate; Fmoc, 9-fluorenylmethoxycarbonyl; HOBt, 1-hydroxybenzotriazole; Mtr, 2,3,6-trimethyl-4-methoxysulfonylbenzene; NOESY, two-dimensional nuclear Overhauser effect spectroscopy; TBTU, 2-(I H-benzotriazol-1 -yl)-1 .I ,3,3-tetramethyluronium tetrafluoroborate; tBu, tert-butyl. benzotriazol-l-yl-oxy-tris(dimethylamino)-phosphonium hexafluorophosphate (BOP) activation and iV-Boc protection [18].A combined method was also described for the (1 -35)-peptide of human neuropeptide Y using solid-phase synthesis by diisopropylcarbodiimide and 1 -hydroxybenzotriazole (HOBt) activation and W-9-fluorenylmethoxycarbonyl (Fmoc) protection followed by enzymatic attachment of the C-terminal tyrosine...

Complete L‐Alanine Scan of Neuropeptide Y Reveals Ligands Binding to Y1 and Y2 Receptors with Distinguished Conformations

“…CD investigations indicate that the Ala substitution at position 34 increases the a-helical content in water, and that no helix stabilization for Ala exchanges in position 35 or even 36 are recorded. According to CD and NMR investigations the existence of two conformations at the C-terminal part comprising residues 33-36 would be possible, a turn-like and a helical conformation (Mierke et al, 1992;Darbon et al, 1992;Li et al, 1992).…”

“…The first conformation of neuropeptide Y resembles a form in which residues 33-36 are arranged in a turn conformation. The second conformation was based upon a NMR study of neuropeptide Y in trifluoroethano1 which indicates an n-helical extension until Gln34 of the C-terminus (Mierke et al, 1992). Starting from these confor- Analytical data (electrospray mass spectra, retention time of HPLC), helicity according to circular dichroism and affinity to SK-N-MC (Y,) and SMS-KAN (Y,) mations molecular dynamics calculations were performed.…”

Are Hormones from the Neuropeptide Y Family Recognized by Their Receptors from the Membrane‐Bound State?

“…In this type of model, the N-terminal tail is fully flexible, whereas residues 11-36 in water [54][55][56] and residues 19-34 in TFE are in an a-helical conformation. [57] Figure 2 displays the conformer bundles derived from the solution structures of PYY and NPY as computed from NMR data. PYY adopts a typical PP-fold-type structure, whereas the N terminus of NPY is fully disordered.…”

Structure–activity relationships of neuropeptide Y analogues with respect to Y1 and Y2 receptors

“…16 However, in the case of each solvent a different number of amino acids participating in the helix was found. A helix composed of amino acids 11-36, 16-36, and 19-34 was identified in water, in dimethylsulfoxide, and in trifluoroethanol, respectively.…”