Improved accuracy of low affinity protein-ligand equilibrium dissociation constants directly determined by electrospray ionization mass spectrometry. - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Journal of The American Society for Mass Spectrometry Année : 2012

Improved accuracy of low affinity protein-ligand equilibrium dissociation constants directly determined by electrospray ionization mass spectrometry.

Résumé

There is continued interest in the determination by ESI-MS of equilibrium dissociation constants (K(D)) that accurately reflect the affinity of a protein-ligand complex in solution. Issues in the measurement of K(D) are compounded in the case of low affinity complexes. Here we present a K(D) measurement method and corresponding mathematical model dealing with both gas-phase dissociation (GPD) and aggregation. To this end, a rational mathematical correction of GPD (f(sat)) is combined with the development of an experimental protocol to deal with gas-phase aggregation. A guide to apply the method to noncovalent protein-ligand systems according to their kinetic behavior is provided. The approach is validated by comparing the K(D) values determined by this method with in-solution K(D) literature values. The influence of the type of molecular interactions and instrumental setup on f(sat) is examined as a first step towards a fine dissection of factors affecting GPD. The method can be reliably applied to a wide array of low affinity systems without the need for a reference ligand or protein.
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Dates et versions

hal-00726223 , version 1 (29-08-2012)

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Citer

Lucie Jaquillard, Fabienne Saab, Françoise Schoentgen, Martine Cadene. Improved accuracy of low affinity protein-ligand equilibrium dissociation constants directly determined by electrospray ionization mass spectrometry.. Journal of The American Society for Mass Spectrometry, 2012, 23 (5), pp.908-22. ⟨10.1007/s13361-011-0305-7⟩. ⟨hal-00726223⟩
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