Development of Inhibitors of UCH-L1 for treatment of neurodegenerative disorders

描述（由申请人提供）：Uch-L1，一种泛素水解酶，与许多神经退行性疾病如阿尔茨海默病（AD）、帕金森病（PD）和亨廷顿病（HD）有关。具体而言，Uch-L1的下调是神经退行性疾病的特征，并且已被证明会降低正常细胞中的突触可塑性，而Uch-L1活性的增加已被证明会增加AD小鼠模型中的突触可塑性和上下文记忆学习。总之，这些数据表明，Uch-L1的催化活性的升高可能是一种有效的方法，在治疗和预防神经退行性疾病。该提议的总体目标是通过X射线晶体学技术表征Uch-L1，以确定可容纳药理学伴侣结合的结合区域，所述药理学伴侣将通过稳定Uch-L1的催化活性构象来增加脑组织中Uch-L1的活性水平，其中药理学伴侣是小分子， 结合并稳定蛋白质，从而增加细胞丰度和活性。期间 在两个具体目标的过程中，我将（1）使用X射线晶体学技术，特别是多溶剂晶体结构（MSCS）方法，鉴定Uch-1_1上能够结合小的药物样分子的非催化结合位点;（2）表征已知增加Uch的催化水解酶活性的单点突变的结构效应，L1，以便开发一种能够模仿突变体效应的伴侣。Petsko-Ringe实验室开发的多溶剂晶体结构（MSCS）方法涉及浸泡在各种溶剂中的酶的X射线晶体学分析。配体结合位点通过检查蛋白质表面的区域来确定，其中多种类型的溶剂分子以高频率结合。在第一个具体目标中，将采用MSCS方法来鉴定Uch-L1的非催化结合位点，基于它们与溶剂分子的相互作用，这些位点可能能够结合药物样分子。

DESCRIPTION (provided by applicant): Uch-L1, an ubiquitin hydrolase, is implicated in numerous neurodegenerative disorders such as Alzheimer's disease (AD), Parkinson's disease (PD), and Huntington's disease (HD). Specifically, down regulation of Uch-L1 is both characteristic of neurodegenerative disorders and has been shown to decrease synaptic plasticity in normal cells, while an increase in Uch-L1 activity has proven to increase synaptic plasticity and contextual memory learning in mouse models of AD. Taken together these data suggest that elevation of the catalytic activity of Uch-L1 may be an effective approach in the treatment and prevention of neurodegenerative disorders. The overall goal of this proposal is to characterize Uch-L1 via X-ray crystallographic techniques to determine binding regions that may accommodate binding of a pharmacological chaperone that will increase levels of the activity of Uch-L1 in brain tissue by stabilization of the catalytically active conformation of Uch-L1, where a pharmacological chaperone is small molecule that binds and stabilizes a protein and, consequently, increase cellular abundance and activity. During the course of the two specific aims I will (1) identify non-catalytic binding sites on Uch-l_1 that are capable of binding small, drug-like molecules using an x-ray crystallography technique, specifically the multiple solvent crystal structure (MSCS) method; (2) characterize the structural effects of a single point mutation that is known to increase the catalytic hydrolase activity of Uch-L1 in order to develop a chaperone that would mimic the effects of the mutant. The multiple solvent crystal structures (MSCS) method developed in the Petsko-Ringe lab involves the x-ray crystallographic analysis of enzymes soaked in various solvents. Ligand binding sites are determined by examination of regions of the protein surface where multiple types of solvent molecules bind with high frequency. In the first specific aim the MSCS approach will be employed to identify non-catalytic binding sites of Uch-L1 that, based on their interactions with solvent molecules, will likely be able to bind drug-like molecules.