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Permanent link (DOI): https://doi.org/10.7939/R3S17T64J

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Screening carbohydrate libraries for protein interactions using the direct ESI-MS assay: Applications to libraries of unknown concentration Open Access

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Author or creator
Kitova, Elena N.
El-Hawiet, Amr
Klassen, John S.
Additional contributors
Subject/Keyword
Carbohydrate
Library screening
Protein interactions
Type of item
Journal Article (Published)
Language
English
Place
Time
Description
A semiquantitative electrospray ionization mass spectrometry (ESI-MS) binding assay suitable for analyzing mixtures of oligosaccharides, at unknown concentrations, for interactions with target proteins is described. The assay relies on the differences in the ratio of the relative abundances of the ligand-bound and free protein ions measured by ESI-MS at two or more initial protein concentrations to distinguish low affinity (≤103 M–1) ligands from moderate and high affinity (>105 M–1) ligands present in the library and to rank their affinities. Control experiments were performed on solutions of a single chain antibody and a mixture of synthetic oligosaccharides, with known affinities, in the absence and presence of a 40-component carbohydrate library to demonstrate the implementation and reliability of the assay. The application of the assay for screening natural libraries of carbohydrates against proteins is also demonstrated using mixtures of human milk oligosaccharides, isolated from breast milk, and fragments of a bacterial toxin and human galectin 3.
Date created
2014
DOI
doi:10.7939/R3S17T64J
License information
© 2014 Kitova, E. N., El-Hawiet, A., & Klassen, J. S. This version of this article is open access and can be downloaded and shared. The original author(s) and source must be cited.
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Citation for previous publication
Kitova, E. N., El-Hawiet, A., & Klassen, J. S. (2014). Screening carbohydrate libraries for protein interactions using the direct ESI-MS assay: Applications to libraries of unknown concentration. Journal of the American Society for Mass Spectrometry, 25(11), 1908-1916.  http://doi.org/10.1007/s13361-014-0964-2

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