SOLUTION STRUCTURE OF HUMAN NEUROPEPTIDE-Y

The three-dimensional structure of synthetic human neuropeptide Y in a queous solution at pH 3.2 and 37 degrees C was determined from two-dim ensional H-1 NMR data recorded at 600 MHz. A restraint set consisting of 440 interproton distance restraints inferred from NOEs and 11 backb one and 4 side-chain dihedral angle restraints derived from spin-spin coupling constants was used as input for distance geometry calculation s in DIANA and simulated annealing and restrained energy minimisation in X-PLOR. The final set of 26 structures is well defined in the regio n of residues 11-36, with a mean pairwise rmsd of 0.51 Angstrom for th e backbone heavy atoms (N, C-alpha and C) and 1.34 A for all heavy ato ms. Residues 13-36 form an amphipathic alpha-helix. The N-terminal 10 residues are poorly defined relative to the helical region, although s ome elements of local structure are apparent. At least one of the thre e prolines in this N-terminal region co-exists in both cis and trans c onformations. An additional set of 24 distances was interpreted as int ermolecular distances within a dimer. A combination of distance geomet ry and restrained simulated annealing yielded a model of the dimer hav ing antiparallel packing of two helical units, whose hydrophobic faces form a well-defined core. Sedimentation equilibrium experiments confi rm the observation that neuropeptide Y associates to form dimers and h igher aggregates under the conditions of the NMR experiments. Our resu lts therefore support the structural features reported for porcine neu ropeptide Y [Cowley, D.J. et al. (1992) Eur. J. Biochem., 205, 1099-11 06] rather than the 'aPP' fold described previously for human neuropep tide Y [Darbon, H. et al. (1992) Eur. J. Biochem., 209, 765-771].