Mass spectrometric analysis of protein–ligand interactions

Abstract

The interactions of small molecules with proteins (protein–ligand interactions) mediate various biological phenomena including signal transduction and protein transcription and translation. Synthetic compounds such as drugs can also bind to target proteins, leading to the inhibition of protein–ligand interactions. These interactions typically accompany association–dissociation equilibrium according to the free energy difference between free and bound states; therefore, the quantitative biophysical analysis of the interactions, which uncovers the stoichiometry and dissociation constant, is important for understanding biological reactions as well as for rational drug development. Mass spectrometry (MS) has been used to determine the precise molecular masses of molecules. Recent advancements in MS enable us to determine the molecular masses of protein–ligand complexes without disrupting the non-covalent interactions through the gentle desolvation of the complexes by in-creasing the vacuum pressure of a chamber in a mass spectrometer. This method is called MS under non-denaturing conditions or native MS and allows the un-ambiguous determination of protein–ligand interactions. Under a few assumptions, MS has also been applied to determine the dissociation constants for protein–ligand interactions. The structural information of a protein– ligand interaction, such as the location of the interaction and conformational change in a protein, can also be ana-lyzed using hydrogen/deuterium exchange MS. In this paper, we briefly describe the history, principle, and recent applications of MS for the study of protein–ligand interactions.