Structure-Based Design of Small Molecules for Aspartylglucosaminuria

描述(申请人提供)：天冬氨酸氨基葡萄糖尿症(AGU)是一种由糖蛋白降解代谢紊乱引起的溶酶体储存性疾病。AgU可导致几乎所有细胞类型的溶酶体中糖天冬酰胺的积聚，并伴有严重的临床症状，如进行性神经变性和智力低下，面部特征粗糙，骨骼异常和结缔组织病变。糖基天冬酰胺酶(GA)的基因发生AgU突变，GA是一种降解糖天冬酰胺所需的溶酶体酶。到目前为止，全世界已经报道了26种不同的AGU等位基因，但仍然没有治疗这种疾病的方法。然而，在过去的几年里，在鉴定和鉴定AGU致病突变方面取得了重大进展。因此，开发一种有效的AGU疗法将使这种遗传性疾病易于对新生儿进行筛查以进行早期治疗。我们对GA的结晶学研究表明，一个表面环(称为前体P-环)阻碍了成熟水解酶的催化中心。因此，为了打开催化中心，需要自体蛋白分解来去除这个P-环。尽管如此，AGU突变会导致GA前体的错误加工和错误定位，从而阻止它们对水解酶活性的自动激活。对GA和AGU分子的高分辨率结构研究将极大地增强我们对这些AGU突变的结构后果的理解，但到目前为止，由于无法获得足够数量的高纯度人类GA，这一研究一直受到阻碍。然而，我们已经克服了这个障碍，通过纯化和结晶细菌GA，它已经被证明具有相同的结构特征，并使用相同的机制来自动激活其水解酶活性。我们的初步结果表明，结构类似甘氨酸的小分子可以增强AGU突变体的自身蛋白分解和水解酶活性。基于我们在GA自动处理方面的最新进展，我们建议在这一应用中1)表征AGU突变的分子发病机制；2)推进GA自身蛋白分解激活的结构和机制研究；3)研究AGU突变的结构后果；以及4)开发小分子来刺激AGU分子的自动处理。这一应用的广泛和长期目标是开发小分子作为治疗药物，以改善AGU误处理和错误定位缺陷，从而减轻AGU患者及其家人的痛苦。

DESCRIPTION (provided by applicant): Aspartylglucosaminuria (AGU) is a lysosomal storage disease caused by a metabolic disorder in glycoprotein degradation. AGU results in accumulation of glycoasparagines in the lysosomes of virtually all cell types, with severe clinical symptoms such as progressive neurodegeneration and mental retardation, coarse facial features, skeletal abnormalities, and connective tissue lesions. AGU mutations occur in the gene for glycosylasparaginase (GA), a lysosomal enzyme required to hydrolyze glycoasparagines. AGU has been reported worldwide, with 26 different AGU alleles found so far, but still no treatment available for this disease. However, during the past few years, there has been significant progress in the identification and characterization of AGU causative mutations. Thus development of an effective AGU therapy would make this genetic disease amenable to newborn screening for an early treatment. Our crystallographic studies on GA reveal that a surface loop (named precursor P-loop) blocks the catalytic center of mature hydrolase. Autoproteolysis is thus required to remove this P-loop in order to open up the catalytic center. Nonetheless, AGU mutations cause misprocessing and mistargeting of GA precursors, thus prevents their autoactivation for the hydrolase activity. High-resolution structural studies of GA and AGU molecules will greatly enhance our understanding of the structural consequences of these AGU mutations but have so far been hampered by the inability to obtain sufficient amounts of highly purified human GA. Nonetheless, we have overcome this hurdle by purifying and crystallizing bacterial GA, which has been demonstrated to have identical structural features and use the same mechanism to autoactivate its hydrolase activity. Our preliminary results indicate that small molecules with structures similar to glycine can enhance autoproteolytic and hydrolase activity of AGU mutants. Building on our recent progress on GA autoprocessing, we propose in this application to 1) characterize molecular pathogenesis of AGU mutations; 2) push forward our structural and mechanistic studies of GA autoproteolytic activation; 3) study structural consequences of AGU mutations; and 4) develop small molecules to stimulate autoprocessing of AGU molecules. The broad, long- term objective of this application is to develop small molecules as therapeutics to ameliorate the AGU misprocessing and mistargeting defect and thus alleviate the suffering of AGU patients and their families.