Resource for Integrated Glycotechnology

描述（由申请人提供）：综合糖技术资源的主要总体目标是开发新的综合技术和多学科方法，以解决糖生物学中的关键问题。蛋白质-碳水化合物相互作用在细胞表面和细胞外基质的许多生物识别事件中起关键作用。我们研究的重点是蛋白聚糖（PG），其中延伸的、线性的、带电荷的聚糖聚合物附着在细胞表面或分泌的蛋白质上，这些蛋白质有助于细胞-细胞、细胞-大分子， 和细胞-基质相互作用，并影响许多生物学功能。蛋白质-PG相互作用的研究是 对蛋白质-碳水化合物相互作用的更广泛的研究。尽管PG在不同的生物过程中至关重要，但关于PG与细胞因子相互作用的细节知之甚少。 结合伴侣或其生物学功能机制。因此，PG-蛋白质相互作用是当代生物学的主要技术前沿。我们将通过利用分析，合成， 结构、生物化学和生物学工具。技术研究和开发（TR&D）项目将应用于几个驱动生物医学项目（DBP）的研究，这些项目将作为综合方法实用性的试验台，并为新的挑战提供反馈，以刺激进一步的技术开发。随着技术的成熟，它们将应用于一系列协作项目，并用于分析服务（C&S）， 并将这些技术的实用性扩大到更广泛的科学界。多种传播战略将提高人们对技术发展的认识和了解，我们的培训课程（D&T）将提供 为研究人员提供直接的机会，从资源人员那里了解最新的技术发展。该提案旨在开发技术（TR& D），通过使用PG片段富集方法、基于MS的测序和组成分析以及化学聚糖低聚物合成和阵列生成进行配体鉴定和验证，推进DBP的目标。PG-蛋白质复合物的结构将使用新的NMR方法和氧化足迹方法来确定，计算方法将预测和验证结构模型。生化目标将产生大量的同位素标记，并进一步修改，以包含顺磁性标签，根据需要，以促进核磁共振研究。含有工程化聚糖聚合物的PG核心蛋白将使用含有增强型或突变型PG生物合成机制的重组表达和细胞系统产生。突变细胞系也将用于表征PG在适当生物学背景下的生物学功能。结合技术的发展将促进我们对这些关键聚糖聚合物结构的理解，并深入了解它们在人类生理学和疾病中的作用。

DESCRIPTION (provided by applicant): The primary Overall objective of the Resource for Integrated Glycotechnology is to develop novel, integrated technologies and multidisciplinary approaches to solve key problems in glycobiology. Protein-carbohydrate interactions play critical roles in numerous biological recognition events at the cell surface and extracellular matrix. The focus of our studies are proteoglycans (PGs), where extended, linear, charged glycan polymers are attached to cell surface or secreted proteins that contribute to cell-cell, cell-macromolecule, and cell-matrix interactions and influence numerous biological functions. Studies on protein-PG interactions are a prototype for more generalized studies on protein-carbohydrate interactions. Despite the critical importance of PGs in diverse biological processes, very little is known about the details of PG interactions with binding partners or their mechanisms of biological function. As a result, PG-protein interactions are a major technical frontier for contemporary biology. We will address the challenges of PG structures, interactions, and biological functions by leveraging advances in analytical, synthetic, structural, biochemical and biological tools. Technology Research and Development (TR&D) projects will be applied to the study of several Driving Biomedical Projects (DBPs) that will act as a test beds for the utility of the integrated approaches and provide feedback for new challenges that will spur further technology development. As the technologies mature they will be applied to a collection of Collaborative Projects and use in Analytical Services (C&S) that will also extend the utility of the technologies to a broader scientific community. Multiple strategies for Dissemination will increase awareness and access to the technology developments and our Training courses (D&T) will provide direct opportunities for researchers to learn the latest technology developments from Resource staff. The proposal aims to develop technologies (TR&Ds) to advance the goals of the DBPs through ligand identification and validation using PG fragment enrichment approaches, MS-based sequencing and composition analysis, and chemical glycan oligomer synthesis and array generation. Structures of PG-protein complexes will be determined using novel NMR methods and oxidative footprinting approaches and computational methods will predict and validate structure models. Biochemical targets will be generated in large quantities with isotopic labels and further modified to contain paramagnetic tags as needed to facilitate NMR studies. PG core proteins containing engineered glycan polymers will be generated using recombinant expression and cell systems harboring enhanced or mutant PG biosynthetic machinery. Mutant cell lines will also be used to characterize biological functions of PGs in appropriate biological contexts. The combined technology developments will advance our understanding of these critical glycan polymer structures and provide insights into their roles in human physiology and disease.