CHARACTERIZATION OF B ANTHRACIS EXOSPORIUM PROTEINS

描述（由申请人提供）：近年来， 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，炭疽病的原因， 大规模杀伤性武器的武库。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，很容易生长， 形成孢子孢子对极端温度的抵抗力， 化学品、干燥和物理损坏使其适合于掺入 用于制造爆炸性武器和隐藏在恐怖主义装置中。孢子可以 通过摄入或吸入进入体内， 细胞，并在1至7天内导致死亡，有时很少或没有明显的 死亡前感染的迹象。抗生素可以用来治疗炭疽， 但是B。炭疽菌株已经或可以被构建成抵抗这些 毒品有一种对抗炭疽毒素的疫苗，但它作用缓慢， 很难生产。因此，更好地应对炭疽威胁是 needed.这项提案的总体目标是确定蛋白质因子对 B的表面。炭疽孢子有助于致病， 新疫苗和药物干预的潜在目标。最外层 是B的。炭疽孢子是一种半透性的、松散的、气球状的 一种叫做外孢壁的结构。大约50%的外孢壁 由一种蛋白质组成，这种蛋白质可能由八种独特的蛋白质成分组成。 目前还没有B的完整外孢壁蛋白。炭疽菌已经 鉴定然而，越来越多的证据表明， 重要的是关键的孢子特性，如结构，活力， 发芽、感染性和毒力。在研究中，我们将 鉴定B中的蛋白质。炭疽孢子外壁，并试图分配一个 每个功能。本建议的三个具体目标是：（1）确定 存在于B的孢子外壁中的蛋白质。炭疽菌和那些 编码这些蛋白质。这将通过分离蛋白质来实现， 质谱法测定部分序列， 以及它们的基因。炭疽基因组;（2） 对编码外孢壁蛋白的基因进行突变， 对上面列出的关键孢子特性的影响;以及（3）使用现有的 一组抗B的单克隆抗体。炭疽孢子表面 (i.e.,外孢壁）以产生亲和力成熟的单链抗体， 孢子外壁蛋白活性，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.