Molecular Probes for Alzheimer's Gamma-Secretase

描述：(申请人提供)淀粉样β蛋白的沉积 (AN)是阿尔茨海默病发病机制中的早期和恒定事件 (AD)，以及从淀粉样前体蛋白中切割Abeta的两种蛋白酶 β-和γ-分泌酶(APP)被认为是重要的治疗靶点。 Y-分泌酶在单个跨膜内催化一种不寻常的蛋白质分解 APP区域，这种切割需要多个跨膜的早老素。 60多种不同的早老素错义突变导致AD，这些 突变改变γ-分泌酶活性以增加高度 纤维蛋白原Abet42残基变体(Abeta42)。抑制剂分析， 分子模拟和诱变研究表明，Y-分泌酶具有 天冬氨酸蛋白酶的性质，并可能催化一种不寻常的膜内 蛋白质分解。此外，早老素中的两个保守的跨膜天冬氨酸 是y-分泌酶活性所必需的，以及过渡态类似物抑制剂。 γ-分泌酶直接和特异地结合早老素。加在一起， 这些结果强烈地表明，早老素本身就是γ-分泌酶， 新型膜内裂解天冬氨酸蛋白酶。当前的目标 建议是了解y-分泌酶活性部位的地形，以 鉴定活性部位内或附近的早老素残基，并 研究γ-分泌酶在体内的正常作用及其化合物的开发 有可能成为阿尔茨海默病的治疗靶点。γ-分泌酶的空间极限 袋S3到S4‘将通过系统地改变 过渡态类似物抑制剂中的相应取代基。这个 强效化合物的发现将使亲和矩阵的发展成为可能 用于该酶的纯化和相关蛋白的鉴定 辅因。与此相互作用的早老素内的特定区域 抑制剂(即活性部位内或附近的部分)将通过 光亲和标记法。黄曲霉毒素跨膜残基的特异性标记 早老素将为膜内活性提供令人信服的证据 地点。最后，这些过渡态模拟物的非肽类似物将是 使用组合化学开发的，作为识别 代谢稳定的化合物，可完全阻断γ-分泌酶活性 有机体。

DESCRIPTION: (Provided by Applicant) Deposition of the amyloid-beta protein (An) is an early and invariant event in the pathogenesis of Alzheimer's disease (AD), and the two proteases that cut Abeta from the amyloid precursor protein (APP), beta- and y-secretases, are considered important therapeutic targets. y-Secretase catalyzes an unusual proteolysis within the single transmembrane region of APP, and this cleavage requires the multi-transmembrane presenilins. More than 60 different missense mutations in presenilins cause AD, and these mutations alter y-secretase activity to increase levels of a highly fibrillogenic 42-residue variant of Abeta (Abeta42). Inhibitor profiling, molecular modeling, and mutagenesis studies reveal that y-secretase has properties of an aspartyl protease and may catalyze an unusual intramembranous proteolysis. Moreover, two conserved transmembrane aspartates in presenilins are required for y-secretase activity, and transition-state analogue inhibitors of y-secretase bind directly and specifically to presenilins. Taken together, these results strongly suggest that presenilins themselves are y-secretases, novel intramembrane-cleaving aspartyl proteases. The objectives of the present proposal are to understand the topography of the y-secretase active site, to identify presenilin residues that are in or near the active site, and to develop compounds for studying the normal role of y-secretase in vivo and its potential as a therapeutic target for AD. The steric limits of y-secretase pockets S3 through S4' will be probed by systematically varying the size of the corresponding substituents in transition-state analogue inhibitors. The discovery of potent compounds would allow the development of affinity matrices for the purification of this protease and identification of associated cofactors. Specific regions within presenilins that interact with such inhibitors (i.e., parts in or near the active site) will be identified by photoaffinity labeling. The specific labeling of transmembrane residues of presenilins would provide compelling evidence for an intramembranous active site. Finally, nonpeptide analogues of these transition-state mimics will be developed using combinatorial chemistry as part of a strategy to identify metabolically stable compounds that block y-secretase activity in whole organisms.