Binding constant determination of high-affinity protein-ligand complexes by electrospray ionization mass spectrometry and ligand competition

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Author(s) Wortmann, Arno, Jecklin, Matthias C., Touboul, David, Badertscher, Martin, Zenobi, Renato
Publication Type Journal Items, Publication Status: Published
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Title Binding constant determination of high-affinity protein-ligand complexes by electrospray ionization mass spectrometry and ligand competition
Author(s) Wortmann, Arno
Jecklin, Matthias C.
Touboul, David
Badertscher, Martin
Zenobi, Renato
Journal or Series Title Journal of mass spectrometry
Volume Number 43
Issue Number 5
Start Page 600
End Page 608
ISSN 1076-5174
1076-5164
Publisher Wiley
Publication Place Chichester
Publication Date 2008
Keyword(s) electrospray ionization mass spectrometry
binding constant determination
high-affinity protein-ligand interaction
competitive ligand binding
avidin
biotin
desthiobiotin
p38 MAP kinase
SB202190
VX-745
Abstract We describe an approach for the determination of binding constants for protein-ligand complexes with electrospray ionization mass spectrometry, based on the observation of unbound ligands competing for binding to a protein target. For the first time, dissociation constants lower than picomolar could be determined with good accuracy by electrospray ionization mass spectrometry. The presented methodology relies only on the determination of signal intensity ratios for free ligands in the low mass region. Therefore, all the advantages of measuring low masses with mass spectrometry, such as high resolution are preserved. By using a reference ligand with known binding affinity, the affinity of a second ligand can be determined. Since no noncovalently bound species are observed, assumptions about response factors are not necessary. The method is validated with ligands binding to avidin and applied to ligands binding to p38 mitogen-activated protein kinase.
DOI 10.1002/jms.1355
Additional Notes Received 16 May 2007, Accepted 26 October 2007, Published online 11 December 2007
Document Type Article
Publication Status Published
Language English
Assigned Organisational Unit(s) 03430
Organisational Unit(s)
NEBIS System Number 001410098
Source Database ID PP-40956
WOS-000256451700005
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@article{Wrtmnn2008,
  author = "Wortmann, Arno and Jecklin, Matthias C. and Touboul, David and Badertscher, Martin and Zenobi, Renato",
  title = "{B}inding constant determination of high-affinity protein-ligand complexes by electrospray ionization mass spectrometry and ligand competition",
  journal = "Journal of mass spectrometry",
  year = 2008,
  volume = "43",
  number = "5",
  pages = "600--608",
}


E-Citations record created: Thu, 01 Apr 2010, 22:43:46 CET