Analysis of three-dimensional structure of ion channel peptides bound to phospholipid bilayers by high resolution solid state NMR

In this research project，have studied to achieve a new approach to determine the three-dimensional structure andmembrane bound ion channel peptides. Following results were obtained in this研究project.(1) The three-dimensional structure of [D113 D1C，-labeled Leu-enkephalin (tyr-Gly-Gly-Phe-Leu) dihydrate crystals was determined on thebasis of six sets of accurately determined - D113 - D1C- D115 - D1N interatomic distances byrotational echo double resonance (REDOR) and some additional constraints from D113 D1C chemicalshifts.(2) isotopically doubly labeled N-Acetyl-Pro-Gly-Phe crystals，obtained from rotational echo double resonance (REDOR) experimentswere analyzed as a three-spin system in which observed nuclei are simultaneously coupled withisotopically labeled intra- and intermolecular heteronuclei. In particular，dipolar interaction was examined to obtain a clue as to the relative orientationof a given molecule to the surrounding neighbors.我们已经开发了一个系统程序determine both the intra- intermolecular interatomic distances and the angle between three twointeratomic vectors.(3) The conformation and dynamics of melittin bound to Thed i米yristoylphophatidylcholin (dmpc) bilayer and the magnetic orientation in the lipid bilayersystems were investigated by solid-state D131 D1P and D113 D1C NMR spectroscopy. UsingD131 D1P NMR，这是发现melitten - dmpc bilayer system forms magnetically oriented elongated vesicles withthe long axis parallel to the magnetic field above the liquid crystalline-gel phase transitionD113 D1C NMR spectra were observed on the D113 D1C labeled melittin bound to thelipid bilayers. Finally，这是发现melittin adopts a transmembrane a-helix whose average axis is parallel to thebilayer normal。

In this research project, we have studied to achieve a new approach to determine the three-dimensional structure and orientation of membrane bound ion channel peptides. Following results were obtained in this research project.(1) The three-dimensional structure of [ィイD113ィエD1C, ィイD115ィエD1N]-labeled Leu-enkephalin (tyr-Gly-Gly-Phe-Leu) dihydrate crystals was determined on the basis of six sets of accurately determined ィイD113ィエD1C-ィイD115ィエD1N interatomic distances by rotational echo double resonance (REDOR) and some additional constraints from ィイD113ィエD1C chemical shifts.(2) Interatomic distances of isotopically doubly labeled N-Acetyl-Pro-Gly-Phe crystals, obtained from rotational echo double resonance (REDOR) experiments, were analyzed as a three-spin system in which observed nuclei are simultaneously coupled with isotopically labeled intra- and intermolecular heteronuclei. In particular, the intermolecular dipolar interaction was examined to obtain a clue as to the relative orientation of a given molecule to the surrounding neighbors. We have developed a systematic procedure to determine both the intra- and intermolecular interatomic distances and the angle between three two interatomic vectors.(3) The conformation and dynamics of melittin bound to the dimyristoylphophatidylcholin (DMPC) bilayer and the magnetic orientation in the lipid bilayer systems were investigated by solid-state ィイD131ィエD1P and ィイD113ィエD1C NMR spectroscopy. Using ィイD131ィエD1P NMR, it was found that melittin-DMPC bilayer system forms magnetically oriented elongated vesicles with the long axis parallel to the magnetic field above the liquid crystalline-gel phase transition temperature. ィイD113ィエD1C NMR spectra were observed on the ィイD113ィエD1C labeled melittin bound to the lipid bilayers. Finally, it was found that melittin adopts a transmembrane a-helix whose average axis is parallel to the bilayer normal.