Structures and lipid interactions of curvature-inducing membrane peptides by NMR

摘要 许多膜上的多肽和蛋白质通过产生膜的曲率来实现它们的功能。 例如，阳离子多肽通过形成永久性的 或瞬时毛孔和病毒融合蛋白，将病毒包膜和靶细胞合并 导致病毒进入的膜。因此，阐明膜的基本机制-- 曲率诱导对于设计无耐药的抗生素和 开发新的疫苗和抗病毒药物。这个项目的长期目标是 了解和量化曲率诱导膜多肽的脂类特异性相互作用。我们 将使用固态核磁共振作为我们的主要工具，因为它是唯一能够同时 探索膜蛋白的高分辨率结构并揭示其物理意义 这些蛋白质所包埋的脂膜的性质。我们提出四个建议 明确的目标。1)我们将研究阳离子多肽在细菌中诱导的脂质聚集- 仿生膜。细菌阴离子和两性离子脂的潜在分离 膜可能是促进膜弯曲的一个重要因素。同位素编辑的核磁共振 将进行实验以测量脂类动力学和多肽-脂类相互作用。 具有代表性的抗菌肽和细胞穿透肽，如PG-1和HIV Tat将是 检查过了。2)我们将开发31P交换核磁共振技术来测量混合料的曲率 脂膜和阳离子多肽在曲率不同区域的定位。3)我们 将决定膜结合的构象、动力学和插入深度 副粘病毒PIV5融合蛋白的融合肽和跨膜区。 PIV5融合蛋白的结构信息将为深入研究PIV5的作用机制提供依据 I类病毒融合蛋白的作用。4)我们将研究PIV5的脂类相互作用 用2H、31P和1H核磁共振仪进行融合多肽实验。膜曲率，多肽 将测量定位和膜水合，以了解多肽是如何修饰的 引起融合的膜结构。

Abstract Many membrane peptides and proteins generate membrane curvature to carry out their function. Examples include cationic peptides that disrupt or cross lipid membranes by forming permanent or transient pores and viral fusion proteins that merge the virus envelope and the target cell membrane to cause virus entry. Thus, elucidating the fundamental mechanism of membrane- curvature induction has broad significance for designing resistance-free antibiotics and for developing new vaccines and antiviral drugs. The long-term objective of this project is to understand and quantify lipid-specific interactions of curvature-inducing membrane peptides. We will use solid-state NMR as our main tool, because it is uniquely capable of simultaneously probing the high-resolution structures of membrane proteins and revealing the physical properties of the lipid membrane in which these proteins are embedded. We propose four specific aims. 1) We will investigate cationic-peptide-induced lipid clustering in bacteria- mimetic membranes. Potential segregation of anionic and zwitterionic lipids of bacterial membranes may be a significant factor in promoting membrane curvature. Isotope-edited NMR experiments will be conducted to measure lipid dynamics and peptide-lipid interactions. Representative antimicrobial and cell-penetrating peptides such as PG-1 and HIV TAT will be examined. 2) We will develop 31P exchange NMR techniques to measure the curvature of mixed lipid membranes and the localization of cationic peptides in curvature-distinct domains. 3) We will determine the membrane-bound conformation, dynamics, and depth of insertion of the fusion peptide and transmembrane domain of the fusion protein of the paramyxovirus, PIV5. Structure information of the PIV5 fusion protein will provide insight into the mechanism of action of class I viral fusion proteins. 4) We will investigate the lipid interactions of the PIV5 fusion peptide using 2H, 31P, and 1H NMR experiments. Membrane curvature, peptide localization and membrane hydration will be measured to understand how the peptide modifies the membrane structure to cause fusion.