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Quantifying labile protein-ligand interactions using electrospray ionization mass spectrometry. | LitMetric

Quantifying labile protein-ligand interactions using electrospray ionization mass spectrometry.

J Am Soc Mass Spectrom

Alberta Ingenuity Centre for Carbohydrate Science, Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada.

Published: November 2010

AI Article Synopsis

Article Abstract

A new electrospray ionization mass spectrometry (ES-MS) approach for quantifying protein-ligand complexes that are prone to in-source (gas-phase) dissociation is described. The method, referred to here as the reference ligand ES-MS method, is based on the direct ES-MS assay and competitive ligand binding. A reference ligand (L(ref)), which binds specifically to the protein (P), at the same binding site as the ligand (L) of interest, with known affinity and forms a stable protein-ligand complex in the gas phase, is added to the solution. The fraction of P bound to L(ref), which is determined directly from the ES mass spectrum, is sensitive to the fraction of P bound to L in solution and enables the affinity of P for L to be determined. A mathematical framework for the implementation of the method in cases where P has one or two specific ligand binding sites is given. Affinities of two carbohydrate-binding proteins, a single chain fragment of a monoclonal antibody and the lectin concanavalin A, for monosaccharide ligands are reported and the results are shown to agree with values obtained using isothermal titration calorimetry.

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Source
http://dx.doi.org/10.1016/j.jasms.2010.07.008DOI Listing

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