X-RAY CRYSTAL STRUCTURE OF CLOSTRIDIUM PERFRINGENS BETA 2 TOXIN

该子项目是利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得主要资金， 因此可以在其他CRISP条目中表示。列出的机构是 中心，不一定是研究者的机构。 β 2毒素是产气荚膜梭菌产生的28 kD外毒素。它与人和动物的坏死性肠炎有关，这是一种以突然急性发作伴致死性出血性粘膜溃疡为特征的疾病。它与牛肉污染有关，导致人类食物中毒。它在宿主细胞膜中形成孔，泄漏必需离子，导致细胞毒性。成孔毒素通常从可溶性单体蛋白转化为形成跨膜通道的寡聚体。膜插入段可以是α螺旋或β折叠的一部分。最近的结构研究提供了一些见解的构象变化与这些蛋白质的膜协会。我们的目标是从结构的角度了解β 2毒素的功能，并确定有助于膜结合，抗原性和孔形成的关键残基。最终，我们希望从解决三维结构中获得的洞察力中开发出乳制品疫苗。 我们用硫酸锂和聚乙二醇3350作为沉淀剂从产气荚膜梭菌中生长了β 2毒素晶体。我们已经研究出了这些晶体的最佳低温条件。在我们当地的X射线源上进行的筛查显示出高达4埃的衍射。我们想 a.收集本地数据集 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