Engineered Organic Color Centers for Profiling Protein-Carbohydrate Recognition

蛋白质-碳水化合物的识别在许多生物过程中都是至关重要的事件，包括细胞间的交流、免疫反应、癌症的发展和转移。由于缺乏通用的探针，了解碳水化合物和碳水化合物结合蛋白之间的特定相互作用一直是一项具有挑战性的任务。在纳米材料主体上产生的有机色心具有精确的光学和碳水化合物功能，特别适合于蛋白质-碳水化合物的识别，这可以导致澄清这两个分子的功能及其潜在的分子机制，并发现治疗和诊断机制。该项目将促进对具有生物功能的工程有机色心结构-性质关系的基本了解，并建立碳水化合物功能化色心在生理和病理途径中对特定生物靶标的探测行为，从而推动科学的进步。除了推进纳米技术、化学、糖科学和工程学的新兴前沿外，这项工作还将对社会福祉产生积极影响。首先，这项工作将有助于开发下一代具有精确功能的多色荧光探针，用于生物医学研究和应用。其次，该项目将通过在不断发展的纳米材料科学与技术和糖科学的跨学科领域通过实践研究促进参与式学习，为所有背景的学生提供令人兴奋的机会。它将进一步支持具有广泛包容性的下一代科学和工程劳动力的发展，特别是在生物纳米技术方面。该项目将专注于通过工程有机色心来探索碳水化合物和蛋白质之间的特定相互作用，目标是实现新的纳米材料工具，以提高灵敏度和选择性来检测生物过程中的目标分子相互作用。建议的色心将通过定向固定化接近天然多糖结构和功能的糖共聚物来共价功能化半导体纯手性单壁碳纳米管的侧壁。建议的颜色中心有许多优点。首先，手性定义的碳纳米管主体具有明确的结构和性质，在纳米管上创建的颜色中心在近红外区域进一步调节光线，减弱了自发荧光和组织深层渗透，为在复杂生物样品中进行高对比度荧光检测提供了理想的条件。第二，纯手性碳纳米管促进了超低剂量、高效率纳米药物的发展。第三，仿生的、精密合成的糖共聚物模拟了碳水化合物在细胞表面的三维显示，因此保证了对蛋白质的更高的灵敏度和选择性。具体地说，拟议的工作将包括：i)使用具有不同碳水化合物配体密度的含半乳糖的糖共聚物合成有机色心；ii)确定半乳糖侧基如何调节色心与人肝癌细胞的去唾液酸糖蛋白受体的特定相互作用；以及iii)评估有机色心的基础毒性和功能性，以利用二维和三维细胞培养选择性地鉴定人肝癌细胞。在这里，细胞培养的微阵列三维生物打印提供了类似活体的微环境，以更好地评估基于纳米材料的颜色中心的毒性和选择性，这些颜色中心可以用作生物兼容的、靶向癌细胞的多色荧光探针。工程有机色心的光学性质及其与蛋白质和人类肝癌细胞的特定相互作用将主要通过色心的光学光谱、高含量成像分析和近红外荧光成像来表征。纳米管结构和糖共聚物构型的组合多样性为化学创新、生化传感和成像技术的发展提供了巨大的潜力。如果成功，这项工作将为研究生物学中的碳水化合物识别提供一种变革性的方法，这对于揭示许多生物过程的分子机制至关重要。这项工作还将创造一种新的多色荧光探针类别，具有以前在许多应用中从未实现的独特特性，如生物传感和生物成像。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Protein-carbohydrate recognitions are crucial events in many biological processes including cell-cell communication, immune response, cancer development and metastasis. Understanding specific interactions between carbohydrates and carbohydrate-binding proteins has been a challenging task due to the lack of versatile probes. Organic color centers created on nanomaterial hosts with precise optical and carbohydrate functionalities are uniquely suited to profile protein-carbohydrate recognition, which can lead to clarifying functions of both molecules and their underlying molecular mechanism and discovering therapeutic and diagnostic mechanism as well. This project will promote the progress in science by advancing fundamental understanding of structure-property relationships of engineered organic color centers with biological functionalities and establishing the probing behavior of carbohydrate-functionalized color centers for specific biological targets in either physiological and pathological pathways. In addition to advancing an emerging frontier across nanotechnology, chemistry, glycoscience, and engineering, this work will positively impact the well-being of society. First, this work will contribute to the development of the next-generation multicolor fluorescent probes with precise functionalities for biomedical research and applications. Second, this project will provide exciting opportunities to students of all backgrounds through promoting engaged learning via hands-on research in an evolving interdisciplinary field of nanomaterial science and technology and glycosciences. It will further support the advancement of a broadly inclusive, next-generation science and engineering workforce, particularly in bio-nanotechnology.This project will focus on probing specific interactions between carbohydrates and proteins by engineered organic color centers with the goal of achieving new nanomaterial tools to detect targeted molecular interactions in biological processes with enhanced sensitivity and selectivity. The proposed color centers will be created through covalently functionalizing the sidewall of semiconducting, pure-chirality single-wall carbon nanotubes via oriented immobilization of glycopolymers that closely mimic the natural glycan structures and functions. There are many advantages for the proposed color centers. First, chirality-defined carbon nanotube hosts have defined structures and properties and color centers created on nanotubes further tune light in the near-infrared regime that has attenuated autofluorescence and deep tissue penetration, providing the ideal condition for high contrast fluorescence detection in complicated biological samples. Second, pure-chirality carbon nanotubes promote advances in ultra-low dose, high efficiency nanomedicines. Third, biomimetic, precision synthesized glycopolymers mimic the three-dimensional display of carbohydrates on the cell surfaces, thus warrant enhanced sensitivity and selectivity for proteins. Particularly, the proposed work will involve i) synthesizing organic color centers using galactose-containing glycopolymers with different carbohydrate ligand densities, ii) determining how the galactose pendent groups of glycopolymers mediate the specific interaction of color centers with asialoglycoprotein receptor of human hepatoma cells, and iii) assessing the basal toxicity and functionality of organic color centers for selectively identifying human hepatoma cells utilizing both two-dimensional and three-dimensional cell cultures. Here, microarray three-dimensional bioprinting of cell cultures provides in vivo-like microenvironments to better asses the toxicity and selectivity of nanomaterials-based color centers, which could be used as biocompatible, cancer cell-targeting multicolor fluorescent probes. Optical properties of engineered organic color centers and their specific interactions with proteins and human liver cancer cells will be characterized primarily by optical spectroscopy of color centers, high-content imaging assays, and near-infrared fluorescence imaging. The combinatorial diversity of nanotube structure and glycopolymer configuration offers vast potential for chemical innovation and biochemical sensing and imaging advancement. If successful, this work will provide a transformative approach to study the carbohydrate recognition in biology that is essential to uncover molecular mechanisms of many biological processes. This work will also open up possibilities to create a new class of multicolor fluorescent probes with distinct properties that are previously never achieved for many applications, such as biosensing and bioimaging.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.