Probing glycosylation in living animals with bioorthogonal chemistries
Jennifer Ann Prescher, University of California, Berkeley (Degree granting institution)
Aberrant glycosylation is a hallmark of numerous disease states. Techniques that enable visualization of glycosylation changes could benefit the development of new diagnostics. Proteins can be readily visualized in living systems using genetically encoded reporters, such as the green fluorescent protein (GFP), or antibodies linked to imaging probes. By contrast, glycans are not amenable to genetically encoded tags, and only a handful of antibodies are known to bind glycans with high affinity. This thesis describes a strategy to equip glycans with reporter tags for visualization and isolation from living systems. As outlined in Chapter 1, the approach relies on the installation of metabolic precursor sugars endowed with unique chemical functionality (e.g., an azide) into target glycans using the cell's own biosynthetic machinery. Azides are non-native functional groups that are inert to biological species, but can be covalently modified via "bioorthogonal" chemical reactions with exogenously delivered probes. Only two such reactions are suitable for use in living systems: the Staudinger ligation of azides with triarylphosphines and the [3 + 2] cycloaddition of azides with cyclooctyne reagents. This two-step labeling process can be used to track glycans in their native environment
Thesis, Dissertation, English, 2006
2006