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

本研究项目旨在从杂交海螺毒液中高通量地鉴定、合成和表征新型神经毒素。以生物多样性为中心的毒素发现策略，加上创新的质谱技术，将用于加速发现新的teretoxin肽序列。然后将鉴定出的teretoxin通过低聚糖-糖基磷脂酰肌醇（GPI）连接物固定在固体载体上，形成teretoxin-GPI微阵列。以微阵列形式接触ter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力学和功能活性的配体。本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技术发现新的地球毒素。2)采用多种化学合成策略合成teretoxin-GPI嵌合体；3)开发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，广泛应用于从蝎子、蛇和蜘蛛以及海蜗牛的巨大肽毒素文库中识别富含半胱氨酸的神经毒素，并开发一种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根本进步。拟议的教育计划将制定和实施一项名为RAISE-W（研究辅助妇女科学赋权计划）的本科实验室研究计划，旨在增加和留住女性，特别是代表性不足的女性，从事STEM学科的科学研究。在STEM领域留住女性的主要障碍是刻板印象和缺乏成功平衡职业/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）确定并招募合格的本科女性，她们将与CUNY的教师配对，从事科学研究项目。2)提供“教练制度”，教授项目参与者时间管理、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.