CHARACTERIZATION OF B ANTHRACIS EXOSPORIUM PROTEINS

描述(由申请人提供)：近年来，越来越多的 外国政府和激进组织，其中许多对美国不友好 及其盟友，已经将炭疽杆菌，炭疽病的起因，添加到他们的 大规模杀伤性武器武器库。这种细菌使用方便。 在战争和恐怖主义中，因为它是高致病性的，可以很容易地生长， 并形成孢子。孢子对极端温度的抵抗力，有毒 化学物质、干燥和物理损害使其适合合并 制造爆炸性武器和藏匿在恐怖分子的装置中。孢子可以 通过摄取或吸入进入体内，萌发成植物人 细胞，并在1至7天内导致死亡，有时很少或没有明显表现 死亡前有感染的迹象。抗生素可以用来治疗炭疽病， 但炭疽杆菌菌株已经或可能被构建成抵抗这些病毒的菌株 毒品。针对炭疽毒素的疫苗是存在的，但它的作用缓慢，而且 很难制作。因此，更好的应对炭疽热威胁的方法是 需要的。这项提案的总体目标是确定蛋白质因素 炭疽芽胞表面对致病有贡献的炭疽芽胞表面 新疫苗和药物干预的潜在目标。最外层 炭疽杆菌的孢子是半透水的、松散的、气球状的。 被称为外孢子菌的结构。大约50%的外孢子菌是 由一种蛋白质组成，该蛋白质可能由八种独特的蛋白质组成。 目前，还没有炭疽杆菌完整的外孢子体蛋白被发现。 已确认身份。然而，越来越多的证据表明，他们将做出贡献 对关键的孢子属性，如结构，活力， 发芽率、传染性和致病力。在拟议的研究中，我们将 鉴定炭疽外孢子菌中的蛋白质，并尝试分配一个 函数传递给每一个。这项建议的三个具体目标是：(1)确定 炭疽芽胞杆菌外孢子中存在的蛋白质和 编码这些蛋白质。这将通过分离蛋白质来完成， 用质谱仪测定部分序列，鉴定蛋白质 以及它们的基因；(2) 突变失活编码外孢子菌蛋白的基因并检测 对上述主要孢子性质的影响；及(3)使用现有的 一组抗炭疽杆菌孢子表面的单抗 (即外孢子菌)产生亲和力成熟的单链抗体，中和 外孢子菌蛋白活性，然后测试这些试剂对孢子性质的影响。

DESCRIPTION (provided by applicant): In recent years a growing number of foreign governments and radical groups, many unfriendly to the United States and its allies, have added Bacillus anthracis, the cause of anthrax, to their arsenals of weapons of mass destruction. This bacterium is convenient for use in warfare and terrorism because it is highly pathogenic, can be grown easily, and forms spores. The spore's resistance to extreme temperatures, noxious chemicals, desiccation, and physical damage make it suitable for incorporation into explosive weapons and for concealment in terrorist devices. Spores can enter the body through ingestion or by inhalation, germinate into vegetative cells, and cause death within 1 to 7 days, sometimes with little or no overt sign of infection preceding death. Antibiotics can be used to treat anthrax, but B. anthracis strains have been, or can be, constructed that resist these drugs. A vaccine against an anthrax toxin exists, but it is slow acting and difficult to produce. Thus, better responses to the threat of anthrax are needed. The overall goal of this proposal is to identify protein factors on the surface of the B. anthracis spore that contribute to pathogenesis and are potential targets for new vaccines and drug intervention. The outermost layer of the B. anthracis spore is a semi-permeable, loose-fitting, balloon-like structure called the exosporium. Approximately 50 percent of the exosporium is composed of a protein consisting of perhaps eight unique protein components. At present, no integral exosporium proteins of B. anthracis have been identified. However, growing evidence suggests that they will contribute significantly to key spore properties such as structure, viability, germination, infectivity, and virulence. In the proposed research, we will identify the proteins in the B. anthracis exosporium and attempt to assign a function to each. The three specific aims of this proposal are: (1) Identify the proteins present in the exosporium of B. anthracis and the genes that encode these proteins. This will be accomplished by isolating proteins, determining partial sequence by mass spectrometry, and identifying proteins and their genes with the aid of the sequenced B. anthracis genome; (2) Mutationally inactivate the genes encoding the exosporium proteins and examine the effects on the key spore properties listed above; and (3) Use an existing panel of monoclonal antibodies against the B. anthracis spore surface (i.e., exosporium) to create affinity-matured single chain antibodies that neutralize exosporium protein activity, then test the effects of these reagents on spore properties.