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

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 在以前利用自旋标记的Gramiidin A(GASL)形成通道的工作中，我们发现在错配的脂类(DPPC，DSPC)的凝胶相中，非通道形式占优势。我们现在已经解决了非通道形式的分配，并研究了甘草苷/脂质体系中的复杂平衡，包括通道(即头对头二聚体[HHD])、非通道二聚体和单体。为了研究可以聚集的自旋标记GA的非通道形式，我们标记了同一分子上的两个不同的位置，而不是测量附着在两个二聚体形成分子上相同位置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。不同分子上的自旋标记物之间的相互作用通过用未标记的标记物基本稀释双自旋标记的标记物来消除。合成并应用了几种新的自旋标记物。在辛醇中，革兰西丁主要以双螺旋左手反平行二聚体的形式存在，我们确定了这种结构的自旋间距。N-末端和C-末端都标记的GAD和N-末端单标记的GALN大大削弱了它们形成HHD的倾向。在大多数脂质中，我们观察到两种不同构象的GAD。主馏分的自旋间距(20.0°)与单体状态一致。双螺旋(31.6°，与辛醇完全相同)是一种次要成分，其含量在饱和脂肪系列中的变化顺序如下：DLPC&lt；DMPC&lt；dSPC&lt；DPPC。DQC还表明，在所研究的所有脂相中，甘草素单体和水解物都只存在于聚集体中。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. 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.