PLIMSTEX: a novel mass spectrometric method for the quantification of protein–ligand interactions in solution

PLIMSTEX: a novel mass spectrometric method for the quantification of protein–ligand... Protein–ligand interactions by mass spectrometry, titration, and H/D exchange (PLIMSTEX) is a new mass spectrometric method for determining association constants and binding stoichiometry for interactions of proteins with various ligands, as well as for quantifying the conformational changes associated with ligand binding to proteins. The association constants determined with PLIMSTEX agree with literature values within a factor of six, establishing its validity for protein interactions involving metal ions, small organic molecules, peptides, and proteins. PLIMSTEX provides solution, not gas-phase, properties by taking advantage of ESI and MALDI mass spectrometry to measure accurately the mass of a protein as it undergoes amide H/D exchange. The approach sidesteps the problem of relating gas-phase abundances of the protein or protein–ligand complex ions to their solution concentrations. With on-column concentration and desalting, high picomole quantities of proteins are sufficient for reproducible mass detection, and the concentration of the protein can be as low as 10 −8 M. It is amenable to different protein/ligand systems in physiologically relevant media. No specially labeled protein or ligand is needed. PLIMSTEX offers minimal perturbation of the binding equilibrium because it uses no denaturants, no additional spectroscopy or reaction probes, and no physical separation of ligand and protein during binding. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png International Journal of Mass Spectrometry Elsevier

PLIMSTEX: a novel mass spectrometric method for the quantification of protein–ligand interactions in solution

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Publisher
Elsevier
Copyright
Copyright © 2004 Elsevier B.V.
ISSN
1387-3806
eISSN
1873-2798
DOI
10.1016/j.ijms.2004.09.012
Publisher site
See Article on Publisher Site

Abstract

Protein–ligand interactions by mass spectrometry, titration, and H/D exchange (PLIMSTEX) is a new mass spectrometric method for determining association constants and binding stoichiometry for interactions of proteins with various ligands, as well as for quantifying the conformational changes associated with ligand binding to proteins. The association constants determined with PLIMSTEX agree with literature values within a factor of six, establishing its validity for protein interactions involving metal ions, small organic molecules, peptides, and proteins. PLIMSTEX provides solution, not gas-phase, properties by taking advantage of ESI and MALDI mass spectrometry to measure accurately the mass of a protein as it undergoes amide H/D exchange. The approach sidesteps the problem of relating gas-phase abundances of the protein or protein–ligand complex ions to their solution concentrations. With on-column concentration and desalting, high picomole quantities of proteins are sufficient for reproducible mass detection, and the concentration of the protein can be as low as 10 −8 M. It is amenable to different protein/ligand systems in physiologically relevant media. No specially labeled protein or ligand is needed. PLIMSTEX offers minimal perturbation of the binding equilibrium because it uses no denaturants, no additional spectroscopy or reaction probes, and no physical separation of ligand and protein during binding.

Journal

International Journal of Mass SpectrometryElsevier

Published: Feb 1, 2005

References

  • Protein–Ligand Interactions
    Metz, G.; Ottleben, H.; Vetter, D.
  • Biochemistry
    Woods, V.L.; Hamuro, Y.
  • Biochem. J.
    Comte, M.; Maulet, Y.; Cox, J.A.

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