UC Berkeley

Glycans are important mediators of many biological processes, including cell-cell adhesion and communication as well as development, inflammation, and cancer cell metastasis. The composition and expression of these biomolecules are known to change during these physiological processes, and, accordingly, there has been much interest in profiling and imaging glycans in vivo. While proteins can be visualized using fluorescent proteins such as GFP, glycans are challenging targets for imaging because they are not genetically encoded. We have developed a two-step method to equip glycans with reporter tags for isolation and visualization from living systems known as the chemical reporter strategy. In our approach, an unnatural sugar, which bears a small reactive functional group known as a chemical reporter, is metabolically incorporated into cell-surface glycans. These reporters can then be detected via a covalent, bioorthogonal reaction such as the Staudinger ligation or Cu-free click chemistry by delivery of exogenous phosphine or cyclooctyne probes, respectively. If an affinity tag or imaging agent is appended to the probe, this method allows for the enrichment and visualization of the labeled glycoconjugates.

This thesis describes the development of new chemical tools for profiling and imaging glycans and their extension to living systems. Chapter 1 provides an overview of recent advances in imaging technologies, including the chemical reporter strategy, for visualizing non-proteinaceous biomolecules such as glycans, lipids, nucleic acids, and small molecule metabolites. Chapter 2 describes the application of Cu-free click chemistry using a panel of cyclooctyne probes to glycan labeling in mice. Chapter 3 describes the synthesis of phosphine and cyclooctyne imaging probes and the evaluation of their ability to image glycans in vivo. Chapter 4 outlines the development of a new chemical reporter for detecting the monosaccharide sialic acid and its application to the labeling of murine glycoconjugates. Finally, Chapter 5 presents a novel strategy for cell-selective labeling of glycans using a caged metabolic precursor.