MEMBRANE-INDUCED SECONDARY STRUCTURES OF NEUROPEPTIDES - A COMPARISONOF THE SOLUTION CONFORMATIONS ADOPTED BY AGONISTS AND ANTAGONISTS OF THE MAMMALIAN TACHYKININ NK1 RECEPTOR

We present what we believe to be the first documented example of an in ducement of distinctly different secondary structure types onto agonis ts and antagonists selective for the same G-coupled protein receptor u sing the same membrane-model matrix wherein the induced structures are consistent with those suggested to be biologically active by extensiv e analogue studies and conventional binding assays. H-1 NMR chemical s hift assignments for the mammalian NK1 receptor-selective agonists alp ha-neurokinin (NKA) and beta-neurokinin (NKB) as well as the mammalian NK1 receptor-selective antagonists [D-Pro(2),D-Phe(7),D-Trp(9)]Sp and [D-Arg(1),D-Pro(2),D-Phe(7),D-His(9)]SP have been determined at 600 M Hz in sodium dodecyl sulfate (SDS) micelles. The SDS micelle system si mulates the membrane-interface environment the peptide experiences whe n in the proximity of the membrane-embedded receptor, allowing for con formational studies that are a rough approximation of in vivo conditio ns. Two-dimensional NMR techniques were used to assign proton resonanc es, and interproton distances were estimated from the observed nuclear Overhauser effects (NOEs). The experimental distances were used as co nstraints in a molecular dynamics and simulated annealing protocol usi ng the modeling package DISCOVER to generate three-dimensional structu res of the two agonists and two antagonists when present in a membrane -model environment to determine possible prebinding ligand conformatio ns. It was determined that (1) NKA is helical from residues 6 to 9, wi th an extended N-terminus; (2) NKB is helical from residues 4 to 10, w ith an extended N-terminus; (3) [D-Pro(2),D-Phe(7),D-Trp(9)]SP has poo rly defined helical properties in the midregion and a beta-turn struct ure in the C-terminus (residues 6-9); and (4) [D-Arg(1),D-Pro(2),D-Phe (7),D-His(9)]SP has a helical structure in the midregion (residues 4-6 ) and a well-defined beta-turn structure in the C-terminus (residues 6 -10). Attempts have been made to correlate the observed conformational differences between the agonists and antagonists to their binding pot encies and biological activity.