Functional Structure of Anti-apoptotic Bcl-2 in Membrane

描述(申请人描述)：细胞凋亡或程序性细胞死亡 消除多细胞生物体中多余或受损的细胞。异常 细胞凋亡在许多人类疾病的发病机制中起作用，包括 癌症。BCL-2及其相关蛋白既可作为抑制子，也可作为启动子 对细胞凋亡的影响。它们的异常表达导致细胞凋亡的失调 以及细胞凋亡性疾病的发生。BCL-2锚定在细胞膜上 线粒体、内质网(ER)或细胞核通过其 C末端跨膜序列。BCL-2维持细胞的正常功能 在许多凋亡事件中的膜，保护细胞器，从而保护 细胞。Bclxl是一种Bcl2同源物，其胞浆结构域的结构如下： 与细菌毒素的成孔区域惊人地相似。尽管 几个Bcl2家族成员在合成的双层膜中形成孔，我们不知道 它们是否在天然膜上形成气孔，如果形成，它们的 成孔活动有助于调节膜的通透性 和细胞凋亡。我们的长期目标是通过以下方式了解分子机制 其中的Bcl2调控细胞凋亡。我们的短期目标是确定 细胞器膜上和细胞膜中具有功能的、天然的Bcl2结构。我们 产生了一系列荧光的全长Bcl-2蛋白，每个蛋白都有一个 附着在特定位置的单一染料。染料标记的Bcl-2蛋白形成 当插入到膜中时，膜孔和光谱会发生变化。因此， Bcl2与膜的相互作用可以用光谱学方法检测和分析。 使用活性的、位点特异性标记的荧光蛋白Bc l-2及其组合 在荧光光谱方法中，我们的目标是表征 BCL-2在内质网微体和线粒体中的表达及其跨膜表达 结构域、孔结构域和胞液结构域。该项目的成果将建立一个 理解Bcl2在调节两者中的功能的结构基础 膜通透性和细胞凋亡，这是设计的先决条件 控制Bcl-2活性的治疗方法，治疗，甚至治愈 凋亡性疾病。

DESCRIPTION (Applicant's Description): Apoptosis or programmed cell death eliminates redundant or damaged cells in multicellular organisms. Abnormal apoptosis contributes to the pathogenesis of many human diseases including cancer. Bcl-2 and related proteins function as either suppressors or promoters of apoptosis. Aberrant expression of them leads to dysregulation of apoptosis and to the onset of apoptotic diseases. Bcl-2 is anchored on the membrane of the mitochondrion, the endoplasmic reticulum (ER) or the nucleus via its C-terminal transmembrane sequence. Bcl-2 maintains the normal function of the membranes in many apoptotic events, protecting the organelles and hence the cells. The structure of the cytosolic domain of Bcl-XL, a Bcl-2 homolog, is strikingly similar to the pore-forming domains of bacterial toxins. Though several Bcl-2 family members form pores in synthetic bilayers, we do not know whether they form pores in the native membrane and if they do, how their pore-forming activities contribute to the regulation of membrane permeability and apoptosis. Our long-term goal is to understand the molecular mechanisms by which Bcl-2 regulates apoptosis. Our short-term goal is to determine the functional, native Bcl-2 structure at and in the organelle membrane. We generated a series of fluorescent, full-length Bcl-2 proteins that each had a single dye attached at a specific site. The dye-labeled Bcl-2 proteins form pores and change spectra upon insertion into membranes. Hence the Bcl-2-membrane interaction can be detected and analyzed spectroscopically. Using the active, site-specifically labeled fluorescent Bcl-2 and a combination of fluorescence spectroscopic methods, we aim to characterize the structure of Bcl-2 at the ER microsomes and the mitochondria with focus on its transmembrane domain, pore and cytosolic domain. The results of the project will establish a structural basis for understanding the functions of Bcl-2 in regulating both membrane permeability and apoptosis, which is a prerequisite for designing therapeutic approaches to control Bcl-2 activity, to treat and perhaps to cure apoptotic diseases.