Year: 2022
Type: article
Abstract: Glycosaminoglycans (GAGs) are a class of highly negatively charged membrane-associated and extracellular matrix polysaccharides involved in the regulation of myriad biological functions, including cell adhesion, migration, signaling, and differentiation, among others. GAGs are typically attached to core proteins, termed proteoglycans (PGs), and can engage >500 binding proteins, making them prominent relays for sensing external stimuli and transducing cellular responses. However, their unique substructural protein-recognition domains that confer their binding specificity remain elusive. While the emergence of glycan arrays has rapidly enabled the profiling of ligand specificities of a range of glycan-binding proteins, their adaptation for the analysis of GAG-binding proteins has been considerably more challenging. Current GAG microarrays primarily employ synthetically defined oligosaccharides, which capture only a fraction of the structural diversity of native GAG polysaccharides. Augmenting existing array platforms to include GAG structures purified from tissues or produced in cells with engineered glycan biosynthetic pathways may significantly advance the understanding of structure-activity relationships in GAG-protein interactions. Here, we demonstrate an efficient and tunable strategy to mimic cellular proteoglycan architectures by conjugating biologically derived GAG chains to a protein scaffold, defined as neoproteoglycans (neoPGs). The use of a reactive fluorogenic linker enabled real-time monitoring of the conjugation reaction efficiency and tuning of the neoPG valency. Immobilization of the reagents on a 96-well array platform allowed for efficient probing of ligand binding and enzyme-substrate specificity, including growth factors and the human sulfatase 1. The neoPGs can also be used directly as soluble probes to evaluate GAG-dependent growth factor signaling in cells.
Source: ACS Chemical Biology
Institution: University of California San Diego
Language: English
Subfield: Cell Biology
Domain: Life Sciences
Open Access status: green
Funders: Alfred P. Sloan Foundation, Research Corporation for Science Advancement, Frederick Gardner Cottrell Foundation, National Institutes of Health, University of California, San Diego, National Heart, Lung, and Blood Institute, National Institute of Child Health and Human Development, Division of Chemistry, Eunice Kennedy Shriver National Institute of Child Health and Human Development
OpenAlex ID: W4280518796