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Related Concept Videos

Electrospray Ionization (ESI) Mass Spectrometry01:12

Electrospray Ionization (ESI) Mass Spectrometry

Higher molecular weight biomolecules are nonvolatile compounds that may decompose before ionizing or vaporizing during mass analysis with conventional electron impact ionization methods. Accordingly, electrospray ionization (ESI) is the favored method for vaporizing and ionizing biomolecules as it circumvents rapid fragmentation and enables the recording of mass signals for the entire biomolecule.
ESI utilizes electrical energy to transfer ions from the liquid phase of the sample into the...
Mass Spectrometry: Overview01:19

Mass Spectrometry: Overview

Mass spectrometry is an analytical technique used to determine the molecular mass and molecular formula of a compound. The basic principle of mass spectrometry is to generate ions from the analyte molecule and measure these ion abundances against their molecular mass. One common type of ionization, known as electron ionization or EI, bombards the analyte molecules in the gas phase with high-energy electron beams. The electron beams displace an electron from the molecule and leave behind a...
Peptide Identification Using Tandem Mass Spectrometry01:33

Peptide Identification Using Tandem Mass Spectrometry

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Related Experiment Video

Updated: Jun 9, 2026

Analyzing Protein Architectures and Protein-Ligand Complexes by Integrative Structural Mass Spectrometry
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Analyzing Protein Architectures and Protein-Ligand Complexes by Integrative Structural Mass Spectrometry

Published on: October 15, 2018

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

Amr El-Hawiet1, Elena N Kitova, Lan Liu

  • 1Alberta Ingenuity Centre for Carbohydrate Science, Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada.

Journal of the American Society for Mass Spectrometry
|August 31, 2010
PubMed
Summary

A novel reference ligand electrospray ionization mass spectrometry (ES-MS) method quantifies protein-ligand binding affinities. This approach overcomes challenges with in-source dissociation, enabling accurate affinity determination for various protein-ligand interactions.

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Last Updated: Jun 9, 2026

Analyzing Protein Architectures and Protein-Ligand Complexes by Integrative Structural Mass Spectrometry
07:33

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Published on: October 15, 2018

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A Protocol for the Identification of Protein-protein Interactions Based on 15N Metabolic Labeling, Immunoprecipitation, Quantitative Mass Spectrometry and Affinity Modulation
14:44

A Protocol for the Identification of Protein-protein Interactions Based on 15N Metabolic Labeling, Immunoprecipitation, Quantitative Mass Spectrometry and Affinity Modulation

Published on: September 24, 2012

Area of Science:

  • Biochemistry
  • Analytical Chemistry
  • Mass Spectrometry

Background:

  • Quantifying protein-ligand interactions is crucial in drug discovery and molecular biology.
  • In-source dissociation during electrospray ionization mass spectrometry (ES-MS) complicates the accurate measurement of these interactions.
  • Existing methods often struggle with protein-ligand complexes prone to gas-phase dissociation.

Purpose of the Study:

  • To develop a robust ES-MS method for quantifying protein-ligand affinities.
  • To address the challenge of in-source dissociation in mass spectrometry-based binding assays.
  • To provide a reliable method for determining the binding affinity of ligands to proteins.

Main Methods:

  • Introduced a reference ligand (L(ref)) with known affinity that forms a stable complex.
  • Utilized competitive ligand binding where L(ref) competes with the ligand of interest (L) for the protein's (P) binding site.
  • Measured the fraction of protein bound to the reference ligand directly from ES-MS spectra.

Main Results:

  • The fraction of protein bound to the reference ligand directly correlates with the fraction bound to the ligand of interest.
  • Developed a mathematical framework for proteins with one or two binding sites.
  • Successfully determined affinities of monosaccharide ligands for two carbohydrate-binding proteins, including concanavalin A.

Conclusions:

  • The reference ligand ES-MS method accurately quantifies protein-ligand affinities, even for complexes prone to dissociation.
  • Results obtained using this method show strong agreement with isothermal titration calorimetry.
  • This technique offers a valuable tool for characterizing biomolecular interactions using mass spectrometry.