Development of a rapid glycosylation profiling system using mass spectrometry dat

描述(申请人提供)：这项提案的最终目标是开发广泛适用的工具来分析复杂基质中的糖肽，并将这些工具应用于母乳和侵袭性乳腺癌细胞的分析。这项工作将通过完成三个具体目标来完成：(1)完成我们的全自动糖肽分析工具包，这是我们在这笔赠款的第一阶段开始的(2)开发 使用大量的包涵物清单来增加MS/MS表征的糖肽离子的数量(3)解决糖肽分析中的两个挑战，这两个挑战超出了当今技术的范围：项目1)识别仅存在于侵袭性乳腺癌中的基于糖肽的生物标记物。项目2)提供了母乳中N-和O-连接的糖肽的完整图谱。大部分工作将通过结合软件设计、生物样本处理、高效液相色谱和质谱学方面的专业知识来开发必要的工具来完成。这些工具的效用将通过将它们应用于完成与女性健康相关的项目来展示：具体地说，我们将提供比以往任何时候都更详细的对母乳独特特性的洞察，我们将表征侵袭性乳腺癌的10-50个已知糖肽标志物上的糖基化。除了我们对妇女健康的贡献外，这项工作还将推动糖蛋白组学领域的发展：糖蛋白分析方法使生物分析技术能够在各种场合使用，从蛋白质结构/功能分析到药物开发，再到生物标记物的发现和开发。

DESCRIPTION (provided by applicant): The ultimate goals of this proposal are to develop broadly applicable tools to analyze glycopeptides in complex matrices and to apply those tools for the analysis of human milk and aggressive breast cancer cells. This work would be accomplished by completing three specific aims: (1) Complete our fully automated glycopeptide analysis tool package, which we started during the first period of this grant's funding (2) Develop the usage of massive inclusion lists to increase the number of glycopeptide ions characterized by MS/MS. (3) Tackle two challenges in glycopeptide analyses that are beyond the reach of today's technology: Project 1) Identify glycopeptide-based biomarkers that are only present in aggressive breast cancer. Project 2) Provide a complete profiling of N- and O-linked glycopeptides in human milk. The bulk of the work would be completed by combining expertise in software design, biological sample handling, HPLC, and mass spectrometry to develop the necessary tools. The utility of the tools would be demonstrated by applying them to complete projects relevant women's health: Specifically, we would provide a more detailed insight into the unique properties of human milk than has ever before been provided, and we would characterize the glycosylation on 10-50 known glycopeptide markers of aggressive breast cancer. In addition to the contributions we make to women's health, this work would drive the field of glycoproteomics forward: The methods for glycoprotein analysis are enabling bioanalytical technologies that can be used in a variety of venues, from protein structure/function analyses to pharmaceutical development, to biomarker discovery and development.