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Permanent link (DOI): https://doi.org/10.7939/R3WS8J00H

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Quantifying protein-fatty acid interactions using electrospray ionization mass spectrometry Open Access

Descriptions

Author or creator
Liu, Lan
Kitova, Elena N.
Klassen, John S.
Additional contributors
Subject/Keyword
Lactoglobulin
Affinity
Fatty acids
Protein-ligand complexes
Electrospray ionization mass spectrometry
Type of item
Journal Article (Published)
Language
English
Place
Time
Description
The application of the direct electrospray ionization mass spectrometry (ESI-MS) assay to quantify interactions between bovine β-lactoglobulin (Lg) and a series of fatty acids (FA), CH3(CH2)xCOOH, where x = 6 (caprylic acid, CpA), 8 (capric acid, CA), 10 (lauric acid, LA), 12 (myristic acid, MA), 14 (palmitic acid, PA) and 16 (stearic acid, SA), is described. Control ESI-MS binding measurements performed on the Lg-PA interaction revealed that both the protonated and deprotonated gas phase ions of the (Lg + PA) complex are prone to dissociate in the ion source, which leads to artificially small association constants (Ka). The addition of imidazole, a stabilizing solution additive, at high concentration (10 mM) increased the relative abundance of (Lg + PA) complex measured by ESI-MS in both positive and negative ion modes. The Ka value measured in negative ion mode and using sampling conditions that minimize in-source dissociation is in good agreement with a value determined using a competitive fluorescence assay. The Ka values measured by ESI-MS for the Lg interactions with MA and SA are also consistent with values expected based on the fluorescence measurements. However, the Ka values measured using optimal sampling conditions in positive ion mode are significantly lower than those measured in negative ion mode for all of the FAs investigated. It is concluded that the protonated gaseous ions of the (Lg + FA) complexes are kinetically less stable than the deprotonated ions. In-source dissociation was significant for the complexes of Lg with the shorter FAs (CpA, CA, and LA) in both modes and, in the case of CpA, no binding could be detected by ESI-MS. The affinities of Lg for CpA, CA, and LA determined using the reference ligand ESI-MS assay, a method for quantifying labile protein–ligand complexes that are prone to in-source dissociation, were found to be in good agreement with reported values.
Date created
2011
DOI
doi:10.7939/R3WS8J00H
License information
© 2011 Liu, L., Kitova, E. N., & 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
Liu, L., Kitova, E. N., & Klassen, J. S. (2011). Quantifying protein-fatty acid interactions using electrospray ionization mass spectrometry. Journal of the American Society for Mass Spectrometry, 22(2), 310-318.  http://doi.org/10.1007/s13361-010-0032-5

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