Capturing Low-Abundance Glycopeptides for Decoding the Glycoproteome

捕获低丰度糖肽以解码糖蛋白组

• 批准号: 10440467
• 负责人: Ronghu Wu
• 金额: $ 29.27万
• 依托单位: GEORGIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-20 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10440467
• 关键词: Acids; Affect; Back; Biological Markers; Biomedical Research; Boronic Acids; Cell Survival; Cells; Clinical; Complex; Covalent Interaction; Cultured Cells; Data; Dendrimers; Detection; Development; Disease; Drug Targeting; Enzymes; Event; Glycopeptides; Glycoproteins; Health; Heterogeneity; Human; Infection; Knowledge; Lead; Life; Malignant Neoplasms; Malignant neoplasm of ovary; Mass Spectrum Analysis; Methods; Mission; Modification; Molecular; Patients; Play; Polysaccharides; Post-Translational Protein Processing; Protein Analysis; Protein Glycosylation; Proteins; Proteomics; Public Health; Research; Role; Sampling; Site; Structure; Surface; Testing; Tissues; United States National Institutes of Health; base; biological research; biological systems; biomarker discovery; cellular development; density; design; disability; early detection biomarkers; glycoproteomics; glycosylation; human disease; hydroxyl group; innovation; insight; monomer; new therapeutic target; novel; operation; potential biomarker; protein function; sugar

SUMMARY Glycosylation is one of the most common protein modifications and is essential for cell survival. Glycoproteins contain a wealth of valuable information regarding the development and disease statuses of cells. Global analysis of protein glycosylation aids in a better understanding of glycoprotein functions and the molecular mechanisms of disease, and leads to the identification of glycoproteins as biomarkers. However, it is extraordinarily challenging to comprehensively analyze glycoproteins because of the heterogeneity of glycans and the low abundance of many glycoproteins. The objective of this project is to develop an innovative and effective method to enrich glycopeptides with diverse glycan structures, especially those with low abundance, and apply this method to globally and site-specifically analyze protein N- and O-glycosylation by mass spectrometry (MS). Guided by strong preliminary data, this objective will be fulfilled by pursuing four specific aims. 1) Effective enrichment of glycopeptides through the synergistic interactions using different types of dendrimers. Based on the common feature that every glycan contains multiple hydroxyl groups, a novel method benefiting from the synergistic interactions between a glycan and multiple boronic acid (BA) molecules conjugated to one dendrimer will be developed to capture low-abundance glycopeptides. Different types of dendrimers will be synthesized and tested, especially from monomers containing the 1→3 branching motif that will increase the density of BA at the dendrimer surface and enhance the interactions with a glycan. 2) Enhancement of the synergistic interactions by minimizing the steric effect and forming the ternary complex. Different kinds of BAs will be studied, especially vinylboronic acids with a small size. This will decrease the steric hindrance and strengthen the overall interaction between one glycan and BAs. Moreover, the formation of the ternary complex will be studied to further enhance the interactions. 3) Global and site-specific analysis of O- glycoproteins with glycan structure information. Through reversible covalent interactions, enriched glycopeptides contain intact glycans, allowing for site-specific analysis of O-glycoproteins with glycan structure information. This is especially important for O-glycosylation due to the lack of an enzyme to universally cleave O-glycans and generate a common tag. 4) Comprehensive analysis of glycoproteins in tissues and sera from patients with ovarian cancer. Combining the proposed method with multiplexed proteomics, glycoproteins in clinical samples will be systematically and quantitatively analyzed. The results will provide insights into the molecular mechanisms of the disease and lead to the discovery of biomarkers for early detection. Eventually, the best dendrimer conjugated with the right BA will enable us to effectively capture low-abundance glycopeptides. Because of the ease of operation and no sample restrictions, the method will have extensive applications in the biological and biomedical research fields and will significantly advance glycoscience.

总结