NON-CHANNEL FORMS OF GRAMICIDIN IN LIPID MEMBRANE BY DQC ESR

通过 DQC ESR 测定脂膜中非通道形式的短杆菌肽

• 批准号: 7956627
• 负责人: BORIS G DZIKOVSKI
• 金额: $ 0.8万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-01 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7956627
• 关键词: Address; Complex; Computer Retrieval of Information on Scientific Projects Database; Equilibrium; Fingerprint; Funding; Gel; Gramicidin; Grant; Hand; Head; Institution; Label; Left; Lipids; Measures; Membrane Lipids; Minor; Molecular Conformation; Octanols; Phase; Positioning Attribute; Research; Research Personnel; Resources; Series; Site; Source; Spin Labels; Structure; System; Technology; United States National Institutes of Health; Work; dimer; gramicidin A; monomer; saturated fat

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. In previous work on channel formation using spin labeled Gramicidin A (GAsl) we found that the non-channel form prevails in the gel phase of mismatching lipids (DPPC, DSPC). We have now addressed the assignment of non-channel forms and studied the complex equilibrium in the gramicidin/lipid system, which includes channels (i.e. head-to-head dimers [HHD]), non-channel dimers, and monomers. To study non-channel forms of spin labeled GA, which can aggregate, we labeled two different sites on the same gramicidin molecule, instead of measuring the distance between spin labels attached at the same position on both dimer-forming molecules. This intramolecular distance between the spin labels could be used as a fingerprint of the particular conformation. Interactions between spin labels on different gramicidin molecules were eliminated by substantially diluting the double spin-labeled gramicidin by unlabeled gramicidin. Several new spin labeled gramicidin compounds were synthesized and used. In octanol, gramicidin exists mainly as a double helical (DH) left-handed antiparallel dimer, and we determined interspin distances for this structure. GAD with both N- and C-termini labeled and GALN, single-labeled at the N-terminus, substantially impair their propensity to HHD formation. In most lipids we observed two different conformations of GAD. The interspin distance in the main fraction (20.0¿) is consistent with the monomeric state. Double helices (31.6¿, exactly as in octanol) are a minor component with the fraction changing in the saturated lipid series in the following order: DLPC<DMPC<DSPC<DPPC. We also showed by DQC that both gramicidin monomers and DH exist in all studied lipid phases only in aggregates.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 在之前使用自旋标记短杆菌肽 A (GAsl) 进行通道形成的工作中，我们发现非通道形式在错配脂质（DPPC、DSPC）的凝胶相中占主导地位。我们现在已经解决了非通道形式的分配问题，并研究了短杆菌肽/脂质系统中的复杂平衡，其中包括通道（即头对头二聚体 [HHD]）、非通道二聚体和单体。为了研究可聚集的自旋标记 GA 的非通道形式，我们在同一短杆菌肽分子上标记了两个不同位点，而不是测量两个二聚体形成分子上相同位置上附着的自旋标记之间的距离。自旋标记之间的分子内距离可以用作特定构象的指纹。通过用未标记的短杆菌肽充分稀释双自旋标记的短杆菌肽，消除了不同短杆菌肽分子上的自旋标记之间的相互作用。合成并使用了几种新的自旋标记短杆菌肽化合物。在辛醇中，短杆菌肽主要以双螺旋（DH）左手反平行二聚体形式存在，我们确定了该结构的自旋间距离。 N 端和 C 端均标记的 GAD 以及 N 端单标记的 GALN 会严重损害其形成 HHD 的倾向。在大多数脂质中，我们观察到 GAD 的两种不同构象。主要部分中的自旋距离（20.0¿）与单体状态一致。双螺旋（31.6¿，与辛醇中完全相同）是次要成分，其分数在饱和脂质系列中按以下顺序变化：DLPC < DMPC < DSPC < DPPC。我们还通过 DQC 表明，短杆菌肽单体和 DH 仅以聚集体形式存在于所有研究的脂质相中。