Influence of Dimehylsulfoxide on Protein–Ligand Binding Affinities

Because of its favorable physicochemical properties, DMSO is the standard solvent for sample storage and handling of compounds in drug discovery. To date, little attention was given to how DMSO influences protein–ligand binding strengths. In this study we investigated the effects of DMSO on different noncovalent protein–ligand complexes, in particular in terms of the binding affinities, which we determined using nanoESI-MS. For the investigation, three different protein–ligand complexes were chosen: trypsin–Pefabloc, lysozyme–tri-N-acetylchitotriose (NAG3), and carbonic anhydrase–chlorothiazide. The DMSO content in the nanoESI buffer was increased systematically from 0.5 to 8%. For all three model systems, it was shown that the binding affinity decreases upon addition of DMSO. Even 0.5–1% DMSO alters the K D values, in particular for the tight binding system carbonic anhydrase–chlorothiazide. The determined dissociation constant (K D) is up to 10 times higher than for a DMSO-free sample in the case of carbonic anhydrase–chlorothiazide binding. For the trypsin–Pefabloc and lysozyme–NAG3 complexes, the dissociation constants are 7 and 3 times larger, respectively, in the presence of DMSO. This work emphasizes the importance of effects of DMSO as a co-solvent for quantification of protein–ligand binding strengths in the early stages of drug discovery.