MOLECULAR BASIS FOR ION CHANNEL FUNCTION

细胞中大约一半的蛋白质与脂质有关。 双层分子，可以说它们执行了一些最有趣和 细胞中的重要功能。例如，离子通道控制 细胞环境和促进神经信号传递，而膜 受体对细胞新陈代谢的控制至关重要。 大分子是病毒感染和免疫的关键要素 Response还吞噬了膜蛋白。不幸的是，相对较少的是 已知的结构，分子操作和组装 膜蛋白。这主要是由于这样一个事实： 可以提供有关水溶性蛋白质的结构信息，例如 结晶学和高分辨率核磁共振在应用于 膜蛋白。当这样的情况下，这可能是一个巨大的回报 结构是被理解的，因为有可能设计出新的 针对这些特定结构和过程的药物 蛋白质。这种结构信息也将有助于分子 对许多遗传性疾病的了解。 目前，还没有电压门控通道，其中分子事件 已经清楚地阐明了导致门控的原因，以及 拟议的研究是为了定义导致 两个膜离子通道的电压门控和调节。该结构 并将研究阿拉米星的门控机制，一种小肽 在膜中产生与电压相关的电导。它是一种 更大的电压门控通道，它也属于更广泛的 具有重要抗生素活性的膜活性多肽。一个 第二种将被研究的蛋白质是磷脂酶。这种蛋白质是 发现于心肌中，是主要的肌膜底物 环磷酸腺苷依赖的蛋白激酶A和蛋白激酶C 是一种8 KDa的跨膜蛋白，形成电压激活的阴离子 频道。通过表征这些分子的结构和门控 渠道，预计有关 膜离子传输的性质及其相关的结构变化 通过选通和通道调节将获得。 拟议的工作将利用光谱技术，如EPR， 高分辨核磁共振和固体核磁共振来确定这些化合物的结构 通道以及与门控和共价修饰相关的变化。 自旋探测器将被结合到这些通道中进行EPR实验 使用合成技术和定点突变。

Approximately half the proteins present in cells are associated with lipid bilayers, and they arguably perform some of the most interesting and important functions in the cell. For example, ion channels control the cellular environment and facilitate nerve signaling, while membrane receptors are critical for the control of cellular metabolism. Macromolecules that are key elements in viral infection and the immune response ate also membrane proteins. Unfortunately, relatively little is known regarding the structures, molecular operation and assembly of membrane proteins. This is primarily a result of the fact that approaches that can yield structural information on water-soluble proteins such as crystallography and high resolution NMR generally fail when applied to membrane proteins. The is potentially an enormous payoff when such structures are understood, because of the potential to design new pharmaceuticals that target specific structures and processes in these proteins. This structural information will also facilitate a molecular understanding of numerous genetic diseases. Presently, there are no voltage-gated channels where the molecular events leading to gating have been clearly elucidated, and the objective of the proposed research is to define the molecular mechanisms that lead to voltage-gating and regulation of two membrane ion channels. The structure and gating mechanism of alamethicin will be studied, a small peptide that produces a voltage-dependent conductance in membranes. It is a model for larger voltage-gated channels, and it also belongs to a wider group of membrane active peptides that have important antibiotic activities. A second protein that will be studied is phospholemman. This protein is found in the myocardium and it is the major sarcolemmal substrate of cyclic AMP dependent protein kinase A and protein kinase C. Phospholemman is an 8 KDa transmembrane protein that forms a voltage-activated anion channel. By characterizing the structure and molecular gating of these channels, it is anticipated that fundamental information regarding the nature of membrane ion transport and the structural changes associated with gating and channel regulation will be obtained. The proposed work will make use of spectroscopic techniques such as EPR, high resolution NMR and solid-state NMR to define the structures of these channels and the changes associated with gating and covalent modification. Spin-probes will be incorporated into these channels for EPR experiments using synthetic techniques and site-directed mutagenesis.