X-RAY CRYSTAL STRUCTURE OF CLOSTRIDIUM PERFRINGENS BETA 2 TOXIN

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 β2毒素是由产气荚膜梭菌产生的一种28kD的外毒素。它与人和动物的坏死性肠炎有关，这种疾病的特征是突然急性发作，并伴有致命的出血性粘膜溃疡。它与牛肉污染有关，导致人类食物中毒。它是在宿主细胞膜上形成孔道，泄漏必需离子，导致细胞毒性。成孔毒素通常从可溶的单体蛋白转变为形成跨膜通道的低聚物。膜插入片段可以是α螺旋或β片状的一部分。最近的结构研究提供了一些关于这些蛋白质在膜结合上的构象变化的洞察力。我们的目标是从结构角度了解β2毒素的功能，并确定有助于膜结合、抗原性和孔道形成的关键残基。最终，我们希望从解决三维结构中获得的洞察力来开发一种乳制品疫苗。 以LiSO4和Peg3350为沉淀剂，从产气荚膜梭菌中培养出β2毒素晶体。我们已经找出了这些晶体的最佳低温条件。在我们本地的X射线源上进行的筛选显示了一些高达4埃的衍射。我们想要 A.收集原生数据集 B.尝试快速卤化物浸泡并收集MAD数据，以便使用MAD技术解决结构问题

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Beta 2 toxin is a 28kD exotoxin produced by Clostridium Perfringens. It is implicated in necrotic enteritis in man and animals, a disease that is characterized by a sudden acute onset with lethal hemorrhagic mucosal ulceration. It is associated with beef contamination resulting in food poisoning in humans. It is pore forming in host cell membranes leaking essential ions leading to cytotoxicity. Pore forming toxins typically transform from soluble monomeric proteins to oligomers that form transmembrane channels. The membrane insertion segment could be alpha helical or part of a beta sheet. Recent structural studies provide some insight into the conformational changes associated with such proteins on membrane association. Our goal is to understand the function of beta 2 toxin from a structural perspective and identify key residues that aid in membrane binding, antigenicity and pore formation. Ultimately we would like to develop a dairy vaccine from the insight gained from solving the 3-dimensional structure. We have grown crystals of beta 2 toxin from Clostridium Perfringens using LiSO4 and Peg3350 as precipitant. We have worked out the best cryo conditions with these crystals. Screening at our local X-ray source has shown some diffraction upto 4 Angstroms. We would like to a. Collect a native data set b. Try quick halide soaks and collect MAD data in order to solve the structure using MAD techniques