STRUCTURAL REGULATION OF CLCA1 ACTIVITY

CLCA1 活动的结构调节

• 批准号: 8706230
• 负责人: THOMAS John BRETT
• 金额: $ 37.24万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-01 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8706230
• 关键词: Address; Affect; Area; Asthma; Automobile Driving; Binding; Biochemical; Biological Assay; Biology; Calcium; Cell model; Cells; Cellular biology; Chloride Channels; Chronic; Chronic Obstructive Airway Disease; Cleaved cell; Consensus; Data; Development; Disease; Drug Targeting; Epithelial Cells; Glycosaminoglycans; Goals; Health; Knowledge; Length; Link; Maintenance; Mediating; Mediator of activation protein; Metalloproteases; Metaplasia; Molecular; Mucous body substance; Pathogenesis; Pathway interactions; Play; Positioning Attribute; Process; Production; Proteins; Proteolysis; Proteolytic Processing; Regulation; Roentgen Rays; Role; Site; Solutions; Structure; Techniques; Therapeutic; Tissues; Translating; Work; X-Ray Crystallography; base; biophysical techniques; design; inhibitor/antagonist; insight; interdisciplinary approach; novel; novel strategies; overexpression; public health relevance; structural biology

DESCRIPTION (provided by applicant): The overall goal of this project is to understand the structural basis of CLCA1 activation and its role in airway biology in health and disease, in order to translate this knowledge into treatments for asthma and COPD, a current unmet need. CLCA1 is a potent modulator of calcium-activated chloride channels (CaCCs) and also a central mediator in mucous cell metaplasia, the process that leads to mucus overproduction. In this project we will investigate the structural and biochemical basis of CLCA activation of CaCCs, and investigate the role that CLCA1-mediated channel activation plays in mucous cell metaplasia. In our preliminary results, we demonstrate that CLCA proteins contain a consensus cleavage site that is recognized by a unique zinicin metalloprotease domain located within the N-terminus of CLCA itself. Furthermore, we show that this self- cleavage is required for hCLCA1 to activate CaCCs. These data suggest that CLCA1 is synthesized in a full- length "inactive" form and that self-cleavage is required to produce an "active" form of the protein. This project will focus on the structural and biochemical analysis of regulation of the metalloprotease domain activity, since it is necessary to produce the active form. We will then characterize the structura changes that occur upon activation by determining the structures of the full-length "inactive" and "active" forms of CLCA1. Finally, we will address the functional role of CLCA1 features in CaCC activation and the role of the channel in mucous cell metaplasia. Understanding how CLCA1 activity is regulated by its own metalloprotease domain and the downstream functional consequences of this regulation will facilitate the design of CLCA1 inhibitors for anti- mucus therapeutics.

描述（由申请人提供）：该项目的总体目标是了解 CLCA1 激活的结构基础及其在健康和疾病中气道生物学中的作用，以便 将这些知识转化为哮喘和慢性阻塞性肺病（目前尚未满足的需求）的治疗方法。 CLCA1 是钙激活氯离子通道 (CaCC) 的有效调节剂，也是粘液细胞化生（导致粘液过量产生的过程）的中心介质。在本项目中，我们将研究 CLCA 激活 CaCC 的结构和生化基础，并研究 CLCA1 介导的通道激活在粘液细胞化生中的作用。在我们的初步结果中，我们证明 CLCA 蛋白含有一个共有切割位点，该切割位点可被位于 CLCA 本身 N 末端的独特锌锌金属蛋白酶结构域识别。此外，我们表明这种自裂解是 hCLCA1 激活 CaCC 所必需的。这些数据表明CLCA1以全长“非活性”形式合成，并且需要自我切割来产生“活性”形式的蛋白质。该项目将重点关注金属蛋白酶结构域活性调节的结构和生化分析，因为它是产生活性形式所必需的。然后，我们将通过确定 CLCA1 全长“非活性”和“活性”形式的结构来表征激活时发生的结构变化。最后，我们将讨论 CLCA1 特征在 CaCC 激活中的功能作用以及该通道在粘液细胞化生中的作用。了解 CLCA1 活性如何受其自身金属蛋白酶结构域调节以及这种调节的下游功能后果将有助于设计用于抗粘液治疗的 CLCA1 抑制剂。