Fibrillogenesis pathways in diabetes and renal diseases

摘要 可溶性蛋白质向淀粉样纤维的转化是许多临床疾病的特征。 包括阿尔茨海默氏症、亨廷顿氏症和II型糖尿病在内的疾病。每例病患的 前体蛋白具有不同的一级、二级和三级结构。但 所得纤维在组织学和超微结构水平上非常相似。纤维 形成动力学类似于结晶，因为存在延长的滞后期 其中纤维是不可检测的。这是其次是合作过渡到纤维 状态有趣的是，正是在滞后阶段采样的中间状态， 被认为是最具细胞毒性的物种。因此，所有这些疾病的核心是 需要确定基础构象变化的分子基础。 这项提案的总体目标是确定淀粉样蛋白的分子基础 两个医学相关系统中的转换。首先，胰岛淀粉样多肽（IAPP）， 由胰腺的胰岛细胞与胰岛素共同分泌的37个残基的肽激素。 在II型糖尿病患者中，IAPP以淀粉样蛋白的形式沉积，并与细胞死亡相关。 第二，需要通过透析治疗的肾脏疾病导致 <$-2微球蛋白（<$2m）淀粉样蛋白在关节引起各种骨骼 病理学在这两种系统中，它都是野生型、未修饰形式的蛋白质 聚集在一起。我们的方法是识别体内环境的变化 并确定这些变化对它们的分子影响。 折叠和原纤维形成。 我们的第一个主要目标是确定构象和寡聚体的变化相关 与IAPP的纤维状组装。重要的是，我们已经确定， IAPP可以被脂质双层以与以下结果一致的方式催化： 糖尿病和肥胖患者的代谢变化。此外，淀粉样蛋白的细胞毒性是 与细胞膜完整性的扰动密切相关。我们将确定 这些效应的分子基础。在200万英镑，我们小组最近发现了一本小说 2 m和Cu（II）之间的相互作用，这可以唯一地引起 在与血液透析期间存在的条件一致的条件下的淀粉样蛋白纤维 疗法我们的第二个主要目标是确定结构和能源基础， 二价诱导的淀粉样变性。我们的目标将通过联合使用诱变， 光学，核磁共振和晶体学技术来阐明蛋白质的扰动 导致纤维形成的结构。 有许多不同的蛋白质，每一种蛋白质在体内都发挥着独特的功能。这些是 由成千上万个原子组成的复杂机器，它们必须折叠成正确的结构， 命令做他们的工作。一组疾病，包括老年痴呆症、糖尿病和肾衰竭 有一个共同的特征，即一种特定的蛋白质错误折叠成纤维结构， 病理这项工作的目的是确定规则管理这种失误使用 2型糖尿病中错误折叠的胰岛淀粉样多肽和微球蛋白 其在用血液透析治疗的肾衰竭患者中错误折叠。

Abstract The conversion of soluble proteins into amyloid fibers is a feature of a number of clinical disorders including Alzheimer¿s, Huntington¿s and type II diabetes. In each case, the precursor protein has distinct primary, secondary and tertiary structure. However, the resultant fibers are remarkably similar at the histological and ultrastructural level. Fiber formation kinetics are similar to crystallization in that there exists a prolonged lag phase in which fiber is undetectable. This is followed by a cooperative transition to the fibrous state. Interestingly, it is the intermediate states sampled during the lag phase that have been identified as the most cytotoxic species. Central to all these disorders, therefore, is the need to identify the molecular basis of the underpinning conformational changes. The overall goal of this proposal is to determine the molecular basis for amyloid conversion in two medically relevant systems. First, islet amyloid polypeptide (IAPP), a 37 residue peptide hormone that is cosecreted with insulin by the ¿-cells of the pancreas. In type II diabetics, IAPP deposits as amyloid and is correlated with ¿-cell death. Second, renal diseases which necessitate treatment by dialysis result in the deposition of ¿-2 microglobulin (¿2m) amyloid in the joints giving rise to a variety of skeletal pathologies. In both of these systems, it is wild-type, unmodified forms of the protein which aggregate. Our approach has been to identify changes in the in vivo environment of theses proteins and to determine the molecular impact of these changes on their folding and fibrillogenesis. Our first major aim is to determine the conformation and oligomeric changes associated with fibrillar assembly of IAPP. Importantly, we have determined that fibrillogenesis of IAPP can be catalyzed by lipid bilayers in a manner consistent with the consequences of metabolic change in diabetics and the obese. Furthermore, cytotoxicity of amyloid is strongly associated with perturbation in cellular membrane integrity. We will determine the molecular basis for these effects. In ¿2m, our group recently discovered a novel interaction between ¿2m and Cu(II) which can uniquely give rise to the nucleation of amyloid fibers under conditions consistent with those present during hemodialysis therapy. Our second major aim is to determine the structural and energetic basis for divalent induced amyloidosis. Our aims will be met by combined use of mutagenesis, optical, NMR and crystallographic techniques to elucidate the perturbation of protein structure that results in fibrillogenesis. There are many different proteins each performing a unique function in the body. These are complex machines composed of thousands of atoms which must fold up to the right structure in order to do their work. A group of diseases, including Alzheimer¿s, diabetes, and renal failure share a common feature in that a particular protein misfolds into fibrous structures that cause pathology. The aim of this work is to determine the rules governing such missteps using the proteins islet amyloid polypeptide, which misfolds in type II diabetes, and ¿-2 microglobulin which misfolds in renal failure patients treated with hemodialysis.