Submicron Emitters Enable Reliable Quantification of Weak Protein-Glycan Interactions by ESI-MS
Erick G Báez Bolivar1, Duong T Bui1, Elena N Kitova1
1Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada T6G 2G2.
Analytical Chemistry
|February 25, 2021
Summary
This study introduces a new method using specialized electrospray ionization mass spectrometry (ESI-MS) to accurately measure weak interactions between carbohydrates (glycans) and glycan-binding proteins (GBPs). This technique overcomes limitations of traditional assays, enabling better understanding of glycan specificity.
Area of Science:
- Biochemistry
- Analytical Chemistry
- Glycobiology
Background:
- Carbohydrate (glycan) and glycan-binding protein (GBP) interactions are crucial for biological processes.
- Weak affinities (dissociation constant > μM) of glycan-GBP complexes hinder specificity determination using conventional assays.
- Accurate measurement of these interactions is essential for understanding biological regulation.
Purpose of the Study:
- To develop a novel ESI-MS method for quantifying low-affinity glycan-GBP interactions.
- To overcome limitations of conventional assays in measuring weak binding affinities.
- To enable direct measurement of glycan specificities for GBPs.
Main Methods:
- Utilized electrospray ionization mass spectrometry (ESI-MS) with nanoflow emitters (∼50 nm inner diameter).
- Minimized nonspecific glycan-GBP binding and adduct formation through submicron emitter technology.
- Performed measurements at physiologically relevant calcium ion concentrations for C-type lectins.
Main Results:
- Successfully quantified low-affinity glycan-GBP interactions (down to ∼5 mM dissociation constant).
- Demonstrated suppression of nonspecific binding and adducts, enabling measurements up to ∼mM glycan concentrations.
- Enabled measurement of calcium-dependent glycan affinities and labile glycan-GBP interactions.
Conclusions:
- Submicron emitter ESI-MS is a powerful tool for characterizing weak glycan-GBP interactions.
- This method facilitates the determination of glycan specificities for GBPs, including C-type lectins.
- The technique advances the study of glycan-mediated biological processes and immune regulation.
