Stabilization of Labile Lysozyme-Ligand Interactions in Native Electrospray Ionization Mass Spectrometry.

Abstract

Flavonoids are polyphenolic secondary metabolites with extensive biological activities and pharmacological effects. Exploring the interactions of flavonoids with proteins may be helpful for understanding their biological processes. Electrospray ionization mass spectrometry (ESI-MS) is a powerful tool to characterize the noncovalent protein-ligand (PL) complexes. However, some protein-flavonoid complexes are labile during electrospray ionization. Here, the labile lysozyme-flavonoid (rutin, icariin, and naringin) complexes were determined by direct ESI-MS without derivation. It has been found that low amounts of N-methylpyrrolidinone and dimethylformamide can protect labile lysozyme-flavonoid complexes away from dissociation during electrospray ionization process. The intact lysozyme-flavonoid complexes were specifically observed in mass spectra, and the measured binding affinities by ESI-MS were matched with the fluorescence data. The effects of additives on the analysis of lysozyme-flavonoid complexes were investigated by ESI-MS, combined with the molecular docking and fluorescence. This strategy was helpful to investigate the labile PL interactions by direct ESI-MS.