CAREER: Development of a teretoxin neuropeptide array for investigating neuronal circuits

这项拟议的研究项目旨在高通量地从陆栖蜗牛的毒液中鉴定、合成和表征新的神经毒素。以生物多样性为中心的毒素发现战略，与创新的质谱学技术相结合，将被用于加快发现新的河豚毒素多肽序列。识别出的毒素将通过固相载体上的寡糖糖基磷脂酰肌醇(GPI)连接物固定，形成Teretoxin-GPI微阵列。以微阵列形式获取毒素的能力有助于发现用于探测神经元回路中离子通道的机制和功能活性的配体。这项研究计划的具体目标是：1)利用电荷增强电子转移解离质谱学技术发现新的毒素。2)采用多种化学合成方法合成Teretoxin-GPI嵌合体；3)研制Teretoxin-GPI神经肽芯片，进行离子通道配体的高通量筛选。这些研究目标的成功完成将产生一种系统的方法，具有广泛的吸引力，可以从蝎子、蛇和蜘蛛以及海洋蜗牛的巨大多肽毒素库中识别富含半胱氨酸的神经毒素，并开发出用于高通量筛选离子通道配体的神经毒素微阵列，这是表征神经元回路的基础进展。拟议的教育计划将开发和实施一项名为RAISE-W(妇女科学赋权研究辅助计划)的本科生实验室研究计划，旨在增加和留住从事STEM学科科学研究的女性，特别是代表性不足的女性的数量。在STEM领域留住妇女的主要障碍是人们认为的陈规定型观念，以及缺乏成功平衡职业/家庭压力的技能。RAISE-W将为克服这些障碍奠定基础。RAISE-W计划的组成部分是：1)在纽约城市大学(CUNY)寻找和招聘合格的本科生女性，她们将与CUNY的教职员工配对从事科学研究项目。2)提供“指导制度”，向项目参与者传授时间管理、成就和职业发展方面的技能。3)通过研究提案奖支持研究生院的活动，确保参与者留在STEM轨道上。

The proposed research project is designed to identify, synthesize, and characterize novel neurotoxins from the venom of terebrid marine snails (teretoxins) in a high throughput manner. A biodiversity centered toxin discovery strategy, coupled with innovative mass spectrometric techniques, will be used to expedite the discovery of novel teretoxin peptide sequences. Identified teretoxins will then be immobilized via an oligosaccharide glycosylphosphatidylinositol (GPI) linker on solid support to form a teretoxin-GPI microarray. The ability to access teretoxins in a microarray format facilitates the discovery of ligands for probing the mechanics and functional activity of ion channels in neuronal circuits. The specific objectives of this research proposal are to: 1) Discover novel teretoxins using charged-enhanced electron-transfer dissociation mass spectrometry techniques. 2) Synthesize teretoxin-GPI chimeras using several chemical synthesis strategies, and 3) Develop a teretoxin-GPI neuropeptide microarray to do high throughput screening of ion channel ligands. The successful completion of these research objectives would produce a systematic approach, with broad appeal for identifying cysteine-rich neurotoxins from the enormous peptide toxin libraries of scorpions, snakes, and spiders, in addition to marine snails, and develop a neurotoxin microarray for high throughput screening of ion channel ligands, a fundamental advance in the characterization of the neuronal circuit. The proposed education initiative will develop and implement an undergraduate laboratory research program termed, RAISE-W (Research Assisted Initiatives in Science Empowerment for Women) that is geared towards increasing and retaining the number of women, especially underrepresented women, engaged in scientific research in STEM disciplines. Key barriers for retaining women in STEM fields are perceived stereotypes and a lack of skills for successfully balancing career/family pressures. RAISE-W will build the foundation for overcoming these obstacles. The components of the RAISE-W program are to: 1) Identify and recruit qualified undergraduate women at The City University of New York (CUNY) who will be paired with CUNY faculty to engage in scientific research projects. 2) Provide a "coaching regime" that teaches program participants skills in time management, achievement, and career development. 3) Support graduate school activities via a research proposal award, ensuring the participants stay on the STEM track.