Collaborative Research: Head Group Preference in Recluse Spider Phospholipase D Toxins

有了这个奖项，化学部的生命过程化学项目正在支持亚利桑那大学的Matthew Cordes博士和刘易斯和克拉克学院的Greta Binford博士之间的合作项目，以确定蜘蛛毒液毒素如何靶向和破坏细胞表面上的不同分子。 棕色隐士蜘蛛的毒液具有导致哺乳动物细胞和组织死亡的毒素，但也有助于蜘蛛捕食和/或消化昆虫猎物。 已知影响哺乳动物的毒素可以通过切断一种叫做胆碱的特定“头基”来破坏细胞表面，这种头基来自一种叫做鞘脂的分子。其他隐士蜘蛛毒素只能切断一种不同的头部基团，称为乙醇胺，哺乳动物的鞘脂中没有这种基团，因此这些毒素对哺乳动物的危害可能较小。许多昆虫都有两种头群，所以这两种类型的毒素可能对捕食都很重要，也许是以不同的方式。科德斯博士是一位结构生物学家和生物化学家，宾福德博士是一位生物学家和有毒无脊椎动物专家，他们联合收割机结合他们的专业知识来确定这些毒素如何区分不同的头部群体，以及对捕食者和猎物的生物后果。这些毒素对不同头部基团的特异性作用也使它们在生物技术中用于检测或操纵不同种类的细胞表面。 该项目的更广泛影响包括通过指导在两个不同机构进行综合和合作研究的本科生进行推广。 蜘蛛引起了公众的兴趣，并提供了一个机会，让公众了解有关这一主题的科学调查。一个被称为“蜘蛛节”的创新项目在实验室和波特兰的一个科学博览会上都得到了展示。本研究的主要目的是了解隐居蜘蛛磷脂酶D毒素中底物头基（乙醇胺与胆碱）偏好的原因和影响。具体目标是映射毒素的界面结合位点，阐明底物头基偏好的氨基酸序列决定因素;表征隐士蜘蛛毒素家族中底物偏好的演变;以及将头基偏好与毒素的生物学效应相关联。使用的方法包括NMR、X射线晶体学、计算结构生物学、定点诱变、系统发育重建、酶测定和生物测定。这项研究的信息阐明了差异识别，蛋白质的作用在膜表面，两个最常见的脂质头组在自然界中，磷酸胆碱和磷酸乙醇胺。该项目还揭示了毒素招聘和专业化毒液和有趣的方面节肢动物生物化学和神经生物学。这些毒素也可以被开发成有价值的分析工具，以探测细胞表面结构的重要差异。该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的知识价值和更广泛的影响审查标准进行评估来支持。

With this award, the Chemistry of Life Processes Program in the Chemistry Division is supporting a collaborative project between Dr. Matthew Cordes at the University of Arizona and Dr. Greta Binford at Lewis and Clark College, to determine how spider venom toxins target and destroy different molecules on cell surfaces. Venoms of brown recluse spiders have toxins that cause the death of cells and tissue in mammals, but also help the spiders immobilize and/or digest insect prey. The toxins that are known to affect mammals can damage cell surfaces by cutting a specific "head group", called choline, off of a molecule called a sphingolipid. Other recluse spider toxins can only cut off a different kind of head group, called ethanolamine, that mammals do not have in their sphingolipids, so these toxins may be less harmful to mammals. Many insects have both kinds of head group, so both types of toxin are probably important for predation, perhaps in different ways. Dr. Cordes, a structural biologist and biochemist, and Dr. Binford, a biologist and expert on venomous invertebrates, combine their expertise to determine how these toxins distinguish the different head groups, and what the biological consequences are for predators and their prey. The specific action of these toxins on different head groups could also make them useful in biotechnology for detecting or manipulating different kinds of cell surfaces. The broader impacts of this project involve outreach through mentorship of undergraduates doing integrative and collaborative research at two different institutions. Spiders attract public interest and afford an opportunity to inform the public on scientific investigations on the subject. An innovative program known as "SpiderFest" is conduucted both at the laboratory and at a science expo in Portland. The overall goal of this project is to understand the causes and effects of substrate head group (ethanolamine vs. choline) preference in phospholipase D toxins from recluse spiders. The specific goals are to map interfacial binding sites in the toxins, elucidate amino-acid sequence determinants of substrate head group preference; characterize the evolution of substrate preference in the recluse spider toxin family; and correlate head group preference to biological effects of the toxins. Methods to be used include NMR, X-ray crystallography, computational structural biology, site-directed mutagenesis, phylogenetic reconstruction, enzymatic assays, and biological assays. Information from this study illuminates differential recognition, by proteins that act at membrane surfaces, of the two most common lipid head groups in nature, phosphocholine and phosphoethanolamine. The project also sheds light on toxin recruitment and specialization in venoms and interesting aspects of arthropod biochemistry and neurobiology. These toxins could also be developed into valuable analytical tools to probe important differences in cell surface structure.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.