Eosinophil Granule Proteins and Their Functions

描述（由申请人提供）：大量证据表明嗜酸性粒细胞在抵抗寄生虫和过敏性疾病（如支气管哮喘）中很重要。嗜酸性粒细胞配备了武器来破坏蠕虫和宿主组织，包括阳离子蛋白、活性氧、白三烯，以及制造细胞因子的能力。在人类疾病中，嗜酸性粒细胞大量积聚并将其颗粒蛋白排放到受影响的组织中，通常与损伤或直接坏死有关。在这项资助下，我们研究了嗜酸性粒细胞颗粒蛋白的结构和功能，包括主要碱性蛋白（MBP1）、嗜酸性粒细胞衍生神经毒素（EDN）、嗜酸性粒细胞阳离子蛋白（ECP）和嗜酸性粒细胞过氧化物酶（EPO）。这些蛋白质是毒素，会损害细胞和组织；MBP1和EPO能够刺激细胞。此外，MBP1会导致猴子和豚鼠的支气管高反应性。我们发现了一种新的颗粒蛋白，即MBP同源物（MBP2）。和MBP1一样，MBP2也会损伤和激活靶细胞。然而，MBP2是否具有MBP1所不具有的独特特性尚不清楚。MBP1作为32 kDa的前分子（proMBP1）合成，并被加工成14 kDa的成熟形式；但人们对proMBP转化酶知之甚少。此外，proMBP1的晶体结构也是未知的。对嗜酸性粒细胞颗粒蛋白质组的初步分析表明存在许多新的肽。在这里，我们确定了一系列相互关联的目标来研究这些问题。首先，我们将比较MBP1和MBP2的属性，确定MBP2是否存在独特的属性。其次，我们将描述将32 kDa的proMBP转化为14 kDa的MBP的酶（proMBP转化酶）。第三，通过合作，我们将对重组proMBP进行结晶并解决其结构问题。第四，我们将利用二维电泳和质谱技术对发现的新型酸性嗜酸性粒细胞颗粒蛋白进行表征。最后，我们将确定在嗜酸性粒细胞增多症患者中是否可以检测到两个新的嗜酸性粒细胞激活标记。总之，这些研究将扩大我们对组成嗜酸性粒细胞颗粒的分子的认识，并为剖析嗜酸性粒细胞在疾病中的功能提供新的工具。

DESCRIPTION (provided by applicant): Considerable evidence indicates that the eosinophil is important in resistance to parasites and in hypersensitivity diseases, such as bronchial asthma. The eosinophil is equipped with weapons to damage helminths and host tissues, including cationic proteins, reactive oxygen species, leukotrienes, and the ability to elaborate cytokines. In human diseases, eosinophils accumulate in massive numbers and discharge their granule proteins onto affected tissues, often in association with damage or frank necrosis. Under the auspices of this grant, we have investigated the structure and function of eosinophil granule proteins, including major basic protein (MBP1), eosinophil-derived neurotoxin (EDN), eosinophil cationic protein (ECP) and eosinophil peroxidase (EPO). These proteins are toxins and damage cells and tissues; MBP1 and EPO are able to stimulate cells. Further, MBP1 causes bronchial hyperreactivity in monkeys and guinea pigs. We have discovered a novel granule protein, the MBP homologue (MBP2). Like MBP1, MBP2 damages and activates target cells. However, whether MBP2 possesses distinctive properties not shared by MBP1 is unknown. MBP1 is synthesized as a 32 kDa promolecule (proMBP1) and is processed to its 14 kDa mature form; but little is known of the proMBP convertase. Also, the crystal structure of proMBP1 is unknown. Preliminary analyses of the eosinophil granule proteome indicate the presence of numerous novel peptides. Here we identify a series of interrelated goals to investigate these issues. First, we will compare the properties of MBP1 and MBP2 and determine whether unique properties exist for MBP2. Second, we will characterize the enzyme(s) converting 32 kDa proMBP to 14 kDa MBP (proMBP convertase). Third, in collaboration, we will crystallize recombinant proMBP and solve its structure. Fourth, we will characterize the novel acidic eosinophil granule proteins discovered using two-dimensional electrophoresis and mass spectroscopy. Finally, we will determine whether two novel markers of eosinophil activation are detectable in patients with eosinophilia. Overall, these studies will expand our knowledge of the molecules composing the eosinophil granule and provide new tools to dissect eosinophil function in disease.