STRUCTURE AND FUNCTION OF PHAGOCYTE PROTEINS

中性粒细胞和其他吞噬血细胞 产生高水平的活性氧物种，以响应 各种感染性或炎症性刺激 呼吸爆发了。这种活性归因于一种酶 一种名为NADPH氧化酶的复合体，它使用分子氧和 NADPH产生超氧阴离子，这是杀微生物剂的前体 氧化剂。这些氧化剂在宿主防御中起着关键作用。 微生物感染并作为炎症信号的中介物。 慢性肉芽肿性疾病患者存在NADPH氧化酶 导致对微生物易感性增加的缺陷 感染和失调的炎症反应。这个项目是 探讨NADPH氧化酶功能的结构基础 呼吸爆发调控的细胞机制。 氧化物酶的激活是一个循序渐进的过程，从 G蛋白偶联受体的刺激和激活 各种激酶和磷脂酶。活性氧化酶组装体 涉及几个蛋白质组分的磷酸化， 转移到特定的膜结构域。我们已经证明了 P47Phox和p67Phox内的相互作用位点(SH3结构域)发挥作用 在调节氧化酶组装中的中心作用，以及这些结构域 既能抑制又能促进活性氧化酶的形成 很复杂。抑制机制包括：1)分子内SH3 P47和p67Phox内的接触，它们维持这些蛋白质在 非活性构象，2)p40Phox的竞争性结合，另 含有SH3结构域的胞浆因子，以及3)一种 RAC激活的激酶，PAK2，通过以下方式下调氧化酶 在磷酸化依赖的过程中结合p47Phox。这个 膜结合的氧化酶复合体的形成是必需的 氧化酶激活，但不充分，因为细胞缺乏 胞浆磷脂酶A2(P85)将组装该氧化酶 络合物，但不产生超氧化物，除非花生四烯酸 如果是这样的话。在相关工作中，我们正在研究 吞噬细胞激活中的磷脂酶D及其产物。 有关NADPH氧化酶的结构和功能的信息可能 为设计治疗策略提供基础，以抑制 或增强呼吸爆发活动。

Neutrophils and other phagocytic blood cells generate high levels of reactive oxygen species in response to a variety of infectious or inflammatory stimuli in a process known as the respiratory burst. This activity is attributed to an enzyme complex called NADPH oxidase, which uses molecular oxygen and NADPH to produce superoxide anion, a precursor of microbicidal oxidants. These oxidants play a key role in host defense against microbial infections and serve as mediators of inflammatory signals. Patients with chronic granulomatous disease have NADPH oxidase deficiencies which result in enhanced susceptibility to microbial infections and dysregulated inflammatory responses. This project is exploring the structural basis for NADPH oxidase function and the cellular mechanisms underlying regulation of the respiratory burst. Oxidase activation is a stepwise process that begins with stimulation of G-protein-coupled receptors and activation of various kinases and phospholipases. Active oxidase assembly involves phosphorylation of several protein components that transmigrate to specific membrane domains. We have shown that interacting sites (SH3 domains) within p47phox and p67phox play a central role in regulating oxidase assembly and that these domains can both inhibit and promote formation of the active oxidase complex. Inhibitory mechanisms include: 1) intramolecular SH3 contacts within p47 and p67phox, which maintain these proteins in inactive conformations, 2) competitive binding of p40phox, another SH3 domain-containing cytosolic factor, and 3) inhibition by a rac-activated kinase, PAK2, which down-regulates the oxidase by binding p47phox in a phosphorylation-dependent process. The formation of the membrane-bound oxidase complex is necessary for oxidase activation, but not sufficient, since cells deficient in cytosolic phospholipase A2 (p85) will assemble the oxidase complex but not produce superoxide unless arachidonic acid is provided. In related work we are examining the role of phospholipase D and its products in phagocyte activation. Information on the structure and function of NADPH oxidase may provide a basis for therapeutic strategies designed to either inhibit or enhance respiratory burst activity.