Functional Structure of Anti-apoptotic Bcl-2 in Membrane

描述（申请人的描述）：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 细胞凋亡有助于许多人类疾病的发病机制， 癌Bcl-2及其相关蛋白质起抑制或促进作用 细胞凋亡。它们的异常表达导致细胞凋亡的失调 以及凋亡性疾病的发病。Bcl-2锚定在细胞膜上， 内质网（ER）或细胞核通过其 C-末端跨膜序列。Bcl-2在维持细胞正常功能的同时， 膜在许多凋亡事件中，保护细胞器，因此， 细胞Bcl-2同源物Bcl-XL的胞质结构域的结构是 与细菌毒素的成孔结构域惊人地相似。虽然 一些Bcl-2家族成员在合成双层中形成孔，我们不知道 它们是否在天然膜上形成孔，如果它们这样做了，它们是如何形成孔的？ 成孔活性有助于调节膜的渗透性 和凋亡。我们的长期目标是了解分子机制， Bcl-2调节细胞凋亡。我们的短期目标是确定 在细胞器膜上和细胞器膜中的功能性天然Bcl-2结构。我们 产生了一系列荧光，全长Bcl-2蛋白，每一个都有一个 附着在特定位点的单一染料。染料标记的Bcl-2蛋白形成 孔并在插入膜中时改变光谱。因此 Bcl-2-膜相互作用可以用光谱法检测和分析。 使用活性的、位点特异性标记的荧光Bcl-2和组合 的荧光光谱方法，我们的目标是表征的结构， Bcl-2在内质网微粒体和线粒体上的表达，重点是其跨膜 结构域、孔和胞质结构域。该项目的成果将建立一个 了解Bcl-2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， 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.