DEVELOPMENT & UTILIZATION OF A MUCIN-GLYCOPEPTIDE MICROARRAY

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心赠款提供。次级项目的主要支助 子项目的主要研究者可能是由其他来源提供的， 包括其它NIH来源。 为子项目列出的总成本可能 表示子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 由于粘蛋白结构域是许多细胞表面分子的组成部分，并且识别这些结构在细胞发育和疾病中很重要，因此我们有兴趣开发用于识别其伴侣和相互作用的有效方法。粘蛋白糖蛋白的基本支架的结构特征在悬垂聚糖表位的展示中是重要的。因此，为了定量表征它们的相互作用，需要相关的糖缀合物构建体而不是分离的聚糖。我们的目标是通过开发一个微阵列平台来满足这一要求，该平台将允许快速筛选合作伙伴。这种结构的可用性一直是这种研究的限制。我们采用了一种基于糖肽化学合成的方法，提供了明确的结构，可以进一步进行生物化学加工。在开发稳健阵列的过程中，正在开发和评估用于将糖肽锚定到载玻片的新型接头和连接方法。分子的初始选择将展示如在肿瘤相关糖蛋白中发现的截短聚糖。 将使用打印结构的单克隆和多克隆抗体评估阵列上分子显示的细节。由我们的化学方法产生的糖缀合物上糖基化的系统变化将允许定义相互作用的选择性。这将扩展到患者血清样本的筛选，以研究抗体癌症生物标志物检测的应用。该项目汇集了格鲁吉亚大学CCRC在化学合成和结构分析方面的专业知识，以及埃默里大学糖组学中心的聚糖相关阵列专业知识。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Because mucin domains are components of a number of cell surface molecules, and recognition of these structures are important in cellular development and disease, we are interested in developing efficient methods for identifying their partners and interactions. Structural characteristics of the underlying scaffold of mucin glycoproteins in the display of pendant glycan epitopes are important. Therefore, to quantitatively characterize their interactions, relevant glycoconjugate constructs are needed rather than isolated glycans. Our goal is to address the requirement through development of a microarray platform that will allow rapid screening for partners. Availability of such constructs has been a limitation in such studies. We have adopted an approach based on glycopeptide chemical synthesis, providing well-defined structures, which can be further elaborated biochemically. In developing a robust array, novel linkers and ligation approaches are being developed and evaluated for anchoring glycopeptides to the slide. The initial selection of molecules will display truncated glycans as found in tumor associated glycoproteins. Details of molecular display on the array will be evaluated using monoclonal and polyclonal antibodies to printed structures. Systematic variation of the glycosylation on the glycoconjugates resulting from our chemical approach will allow definition of the selectivity of the interactions. This will be extended to screening of patient serum samples to investigate applications to detection of antibody cancer biomarkers. The project brings together the expertise in chemical synthesis and structural analysis at the CCRC at the University of Georgia, and glycan related array expertise at the Glycomics Center at Emory University.