# Effective Methods to Globally Analyze Cell Surface Proteins and Glycoproteins > **NIH NIH R01** · GEORGIA INSTITUTE OF TECHNOLOGY · 2020 · $343,156 ## Abstract SUMMARY: Cell surface proteins and glycoproteins are essential for mammalian cell survival, and they frequently modulate and represent different developmental and diseased statuses of cells. Many important surface proteins, such as receptors, transporters and enzymes, are of extraordinary clinical interest as therapeutic targets, and they can also serve as valuable biomarkers for disease detection. Despite the critical importance of surface proteins and glycoproteins, there is a substantial gap between their global analysis and effective methods available to achieve it. The objective of this project is to globally analyze surface proteins and glycoproteins by designing innovative and effective methods integrating click chemistry, enzymatic reactions and mass spectrometry- based proteomics. Guided by strong preliminary data, this objective will be fulfilled by pursuing three specific aims. (1) Develop an effective method for the comprehensive analysis of surface proteins. Surface proteins in living cells will be efficiently tagged via copper-free click chemistry. After cell lysis, surface proteins with the specific tag will be covalently bound to beads via a second well-designed click reaction. The covalent connections will allow the complete removal of non-specifically bound proteins by denaturing enriched surface proteins, for specific and global surface protein identification. (2) Identify N-glycoproteins on the cell surface globally and site-specifically. New methods integrating mild enzymatic and chemical reactions will be designed to target surface glycoproteins, including proteins bearing a particular glycan, for MS analysis. (3) Study surface protein secretion and systematically quantify surface proteins and glycoproteins in cancer cells. Novel methods will provide a unique opportunity to investigate surface proteins and glycoproteins in cancer cells with progressive invasiveness. The proposed research contains five innovations to decode the cell surface proteome and glycoproteome. First, cell surface proteins will be tagged through a copper-free click chemistry reaction for the first time, which is ideal due to its high speed, specificity and mild reaction conditions. Second, covalent interactions will be employed to connect surface proteins containing the specific tag with magnetic beads. This will enable the complete removal of non-specifically bound proteins, and overcome the long- standing obstacle. Third, surface protein dynamics will be systematically studied and their half-lives will be measured. Fourth, comprehensive and site-specific identification of surface glycoproteins will be achieved, and surface glycoproteins containing a particular glycan will be selectively analyzed. Fifth, surface protein secretion will be globally investigated, and surface proteins and glycoproteins will be quantified among different cancer cells with progressive invasiveness. These methods will have extensive applications in biomedical research by pro... ## Key facts - **NIH application ID:** 9851406 - **Project number:** 5R01GM118803-04 - **Recipient organization:** GEORGIA INSTITUTE OF TECHNOLOGY - **Principal Investigator:** Ronghu Wu - **Activity code:** R01 (R01, R21, SBIR, etc.) - **Funding institute:** NIH - **Fiscal year:** 2020 - **Award amount:** $343,156 - **Award type:** 5 - **Project period:** 2017-02-01 → 2022-07-31 ## Primary source NIH RePORTER: https://reporter.nih.gov/project-details/9851406 ## Citation > US National Institutes of Health, RePORTER application 9851406, Effective Methods to Globally Analyze Cell Surface Proteins and Glycoproteins (5R01GM118803-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9851406. Licensed CC0. --- *[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*