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

描述(申请人提供)：UCH-L1是一种泛素水解酶，与许多神经退行性疾病有关，如阿尔茨海默病(AD)、帕金森病(PD)和亨廷顿病(HD)。具体地说，Uch-L1的下调既是神经退行性疾病的特征，也被证明降低了正常细胞的突触可塑性，而Uch-L1活性的增加被证明增加了AD模型小鼠的突触可塑性和上下文记忆学习。综上所述，这些数据表明，提高Uch-L1的催化活性可能是治疗和预防神经退行性疾病的有效方法。该提案的总体目标是通过X射线结晶学技术表征Uch-L1，以确定可能与药理伴侣结合的结合区域，该结合区域将通过稳定Uch-L1的催化活性构象来增加Uch-L1在脑组织中的活性水平，其中药理伴侣是指 结合并稳定蛋白质，从而增加细胞的丰富性和活性。在.期间 在这两个特定目标的过程中，我将(1)利用X射线结晶学技术，特别是多溶剂晶体结构(MSCS)方法，确定Uch-L_1上能够结合小分子类药物的非催化结合位点；(2)表征已知可提高Uch-L1的催化水解酶活性的单点突变的结构效应，以便开发出一种模仿突变体影响的伴侣。Petsko-Ringer实验室开发的多溶剂晶体结构(MSCS)方法包括对浸泡在各种溶剂中的酶进行X射线结晶学分析。配体结合位置是通过检查蛋白质表面的区域来确定的，在这些区域中，多种类型的溶剂分子频繁结合。在第一个具体目标中，将使用MSCs方法来确定Uch-L1的非催化结合部位，基于它们与溶剂分子的相互作用，这些结合部位很可能能够结合类药物分子。

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.