Dual Oxidases (Duox): Enzymology & Biological Function

双氧化酶 (Duox)：酶学

• 批准号: 6742511
• 负责人: John David Lambeth
• 金额: $ 26.75万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-05-01 至 2007-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6742511
• 关键词: Drosophilidae; NAD(P)H dehydrogenase; biological models; calcium; cell membrane; clinical research; crosslink; enzyme activity; enzyme mechanism; enzyme structure; epithelium; extracellular matrix; hormone biosynthesis; human genetic material tag; human tissue; immunity; immunofluorescence technique; iodotyrosine; mass spectrometry; oxidation; peroxidases; phage display; protein localization; thyroid hormones; tissue /cell culture; tyrosine

DESCRIPTION (provided by applicant): A family of Dual Oxidases (Duox) has been defined. Duox enzymes consist of three domains: 1) an NADPH-oxidase domain homologous to gp91phox, the catalytic subunit of the phagocyte respiratory burst oxidase; 2) a calcium-binding domain; and 3) a peroxidase domain. Located in plasma membrane of epithelial cells (colon, lung, thyroid, pancreas), Duoxs' are proposed to function by using intracellular NADPH to reduce 02 to form H202 outside the cell. The latter is used by the peroxidase domain to catalyze peroxidative reactions involving modification of extracellular matrix proteins and probably other extracellular small molecules. Specifically, Duox's are proposed to function biologically in innate immunity, endocrine function and cancer. We identified and cloned two human isoforms, h-Duox1 and h-Duox2, and have identified Duox enzymes in diverse species, including C. elegans and Drosophila. In C. elegans, we showed that the enzyme functions to chemically cross-link tyrosine residues in collagen and other extracellular matrix proteins, thus stabilizing the structure of the cuticle. We will test the hypothesis that a general function of Duox enzymes throughout the animal kingdom is the chemical modification of tyrosine and/or other residues of extracellular matrix. Genetic and biochemical methods will be used to test this hypothesis in Drosophila, where we propose that the tyrosine modifications stabilize the structure of the wing. Phage display methods will be used to develop peptide inhibitors of the peroxidase domains of h-Duoxl and h-Duox2, and these will be used to investigate normal biological functions of Duox such as thyroid hormone biosynthesis and innate immunity. Enzymatic properties and regulation by calcium of Duox will be documented, and the transmembrane topology of the domains will be explored. These studies will provide for the first time fundamental information regarding the enzymology, topology and biological functions of this newly discovered group of enzymes and will document their ability to chemically modify extracellular matrix (ECM). Because ECM is a critical determinant of transformation Duox enzymes may play an important role in cancer biology in some tissues.

描述（由申请人提供）：双氧化酶家族（Duox）已被定义。Duox酶由三个结构域组成：1)与吞噬细胞呼吸爆发氧化酶的催化亚基gp91phox同源的nadph氧化酶结构域；2)钙结合域；3)过氧化物酶结构域。Duoxs'位于上皮细胞（结肠、肺、甲状腺、胰腺）的质膜上，通过细胞内NADPH还原02在细胞外形成H202发挥作用。后者被过氧化物酶结构域用于催化涉及细胞外基质蛋白和其他细胞外小分子修饰的过氧化反应。具体来说，Duox被认为在先天免疫、内分泌功能和癌症中发挥生物学作用。我们鉴定并克隆了两种人类同种异构体，h-Duox1和h-Duox2，并在多种物种中鉴定了Duox酶，包括秀丽隐杆线虫和果蝇。在秀丽隐杆线虫中，我们发现该酶可以化学交联胶原蛋白和其他细胞外基质蛋白中的酪氨酸残基，从而稳定角质层的结构。我们将验证这个假设，即在整个动物王国中，Duox酶的一般功能是对酪氨酸和/或细胞外基质的其他残基进行化学修饰。遗传和生化方法将用于在果蝇中测试这一假设，我们提出酪氨酸修饰稳定了翅膀的结构。噬菌体展示方法将用于开发h-Duoxl和h-Duox2过氧化物酶结构域的肽抑制剂，这些抑制剂将用于研究Duox的正常生物学功能，如甲状腺激素的生物合成和先天免疫。酶的性质和调节的氧化钙将被记录，并跨膜拓扑结构域将被探索。这些研究将首次提供有关这组新发现酶的酶学、拓扑结构和生物学功能的基本信息，并将记录它们化学修饰细胞外基质（ECM）的能力。由于ECM是转化的关键决定因素，在某些组织中，Duox酶可能在癌症生物学中发挥重要作用。