A metabolically engineered spin-labeling approach for studying glycans on cells

Area of Science:

• Biochemistry
• Cell Biology
• Biophysics

Background:

• Cell surface glycans form a complex and heterogeneous layer, the glycocalyx, crucial for cellular functions.
• Understanding the local environment and organization of glycans is vital for deciphering cellular processes and disease states.

Purpose of the Study:

• To investigate the heterogeneous microenvironment of cell surface glycans in cancer and normal cells using metabolic glycan engineering and spin-labeling.
• To differentiate the packing and mobility of specific glycan moieties, such as sialic acid and N-acetylglucosamine (GlcNAc).

Main Methods:

• Metabolic glycan engineering (MGE) to incorporate azide groups into cell surface glycans.
• Nitroxide spin-labeling (SL) via click chemistry to attach spin labels to azide-modified glycans.
• Electron Paramagnetic Resonance (EPR) spectroscopy to analyze the mobility and microenvironment of labeled glycans.

Main Results:

• Both sialic acid and GlcNAc exhibited heterogeneous environments, characterized by a combination of mobile and restricted spectral components.
• Sialic acid moieties were found in a less crowded environment (∼80% more mobile), while GlcNAc sites were in a more crowded environment (∼50% more restricted).
• Distinct cell lines displayed varied proportions of these components, indicating differences in glycan organization and composition.

Conclusions:

• The study demonstrates the utility of spin-labeling electron paramagnetic resonance (SL-EPR) for studying cellular glycans.
• SL-EPR provides insights into the local organization and heterogeneity of the cellular glycocalyx.
• This approach can distinguish differences in glycan packing and composition among various cell types.