title: Quantifying protein-carbohydrate interactions using liquid sample desorption electrospray ionization mass spectrometry.
abstract: The application of liquid sample desorption electrospray ionization mass spectrometry (liquid sample DESI-MS) for quantifying protein-carbohydrate interactions in vitro is described. Association constants for the interactions between lysozyme and β-D-GlcNAc-(1 → 4)-β-D-GlcNAc-(1 → 4)-D-GlcNAc and β-D-GlcNAc-(1 → 4)-β-D-GlcNAc-(1 → 4)-β-D-GlcNAc-(1 → 4)-D-GlcNAc, and between a single chain antibody and α-D-Galp-(1 → 2)-[α-D-Abep-(1 → 3)]-α-D-Manp-OCH3 and β-D-Glcp-(1 → 2)-[α-D-Abep-(1 → 3)]-α-D-Manp-OCH3 measured using liquid sample DESI-MS were found to be in good agreement with values measured by isothermal titration calorimetry and the direct ESI-MS assay. The reference protein method, which was originally developed to correct ESI mass spectra for the occurrence of nonspecific ligand-protein binding, was shown to reliably correct liquid sample DESI mass spectra for nonspecific binding. The suitability of liquid sample DESI-MS for quantitative binding measurements carried out using solutions containing high concentrations of the nonvolatile biological buffer phosphate buffered saline (PBS) was also explored. Binding of lysozyme to β-D-GlcNAc-(1 → 4)-β-D-GlcNAc-(1 → 4)-D-GlcNAc in aqueous solutions containing up to 1× PBS was successfully monitored using liquid sample DESI-MS; with ESI-MS the binding measurements were limited to concentrations less than 0.02 X PBS.
authors.affiliation: Alberta Glycomics Center and Department of Chemistry, University of Alberta, Edmonton, AB, T6G 2G2, Canada.
authors.forename: Yuyu
authors.initials: Y
authors.lastname: Yao
authors.forename: Km
authors.initials: K
authors.lastname: Shams-Ud-Doha
authors.forename: Rambod
authors.initials: R
authors.lastname: Daneshfar
authors.forename: Elena N
authors.initials: EN
authors.lastname: Kitova
authors.forename: John S
authors.initials: JS
authors.lastname: Klassen
mesh_terms: D002241:Carbohydrates / Q000737:chemistry*; D008956:Models, Chemical; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D012965:Sodium Chloride; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
chemical_list: D002241:Carbohydrates; D011506:Proteins; D012965:Sodium Chloride