Preliminary studies of C. botulinum progenitor toxin B

描述(申请人提供)：肉毒杆菌神经毒素是一种由肉毒梭菌释放的称为前体毒素的蛋白质复合体，如果不立即和适当的治疗，会导致肉毒杆菌中毒，因为它的死亡率很高。这也是一个潜在的生物武器威胁。它可能用于生物恐怖主义，因此必须制定保护武装部队和一般公众的措施。这项提案的总体目标是帮助制定和设计此类措施，即药品和疫苗。长期目标是通过了解肉毒弧菌前体毒素(由神经毒素和其他相关蛋白质形成的复合体)的组成成分组装的功能和目的，通过X射线结晶学确定前体毒素B的三维结构来实现这一目标。据信，前体毒素保护肉毒杆菌神经毒素免受胃肠道中存在的低pH和肽酶的恶劣条件的影响。了解这些蛋白质的三维结构以及它们与神经毒素的相互作用将有助于理解它们保护神经毒素的机制。前体毒素能够以最低限度的降解从肠道吸收，通过适当修饰毒素分子，使其适合用作口服药物的载体。此外，它还将有助于设计口服疫苗，以治疗肉毒杆菌中毒和破伤风以外的许多疾病。当包括动物和鸟类在内的大量人口要接种疫苗时，这一点尤其重要。短期目标是使M型和L型前体毒素B结晶，用于X射线衍射分析，并在获得晶体后启动结构分析。通向总体目标的个人具体目标如下所示。1.将肉毒杆菌前体毒素B的M型和L型结晶，用于X射线结晶学三维结构测定。2.利用布鲁克黑文国家实验室的STEM设备测量祖细胞复合体的质量。3.从M型和L型前体毒素B的晶体中获得初步的晶体数据。

DESCRIPTION (provided by applicant): Botutinum neurotoxin, released as a protein complex called progenitor toxin by the bacterium Clostridium botulinum, causes the disease botulism that is of considerable concern because of its high mortality rate when not treated immediately and properly. It is also a potential biowarfare threat. Its potential use in bioterrorism makes it imperative to develop measures to protect the armed forces and the public in general. The overall objective of this proposal is to aid in developing and designing such measures viz., medicines and vaccines. The long-range goal is to achieve this by understanding the function and purpose of the assembly of the components of C. botutinum progenitor toxins, complex formed by neurotoxin and other associated proteins, by determining the three-dimensional structure of progenitor toxin B via X-ray crystallography. It is believed that progenitor toxins protect the botulinum neurotoxin from the harsh conditions of low pH and peptidases present in the gastrointestinal tract. Knowledge of the three-dimensional organization of these proteins and their interactions with the neurotoxin and one another will lead to the understanding of the mechanism by which they protect the neurotoxin. Progenitor toxin's ability to be absorbed from the gut with minimum degradation makes it a suitable candidate for use as a carrier of oral medicines by appropriate modification of the toxin molecule. Additionally, it will help in designing oral vaccines for many afflictions besides botulism and tetanus. This is particularly important when a large population including animals and birds is to be vaccinated. The short-range goals are to crystallize M and L forms of progenitor toxin B for X-ray diffraction analysis and initiate structural analysis when crystals are obtained. The individual specific aims leading to the overall goal are listed below. 1. To crystallize M and L forms of C. botulinum progenitor toxin B for three-dimensional structure determination via X-ray crystallography. 2. To measure the mass of the progenitor complex with the use of the STEM facility at Brookhaven National Laboratory. 3. To obtain preliminary crystal data from the crystals of M and L forms of progenitor toxin B.