Probing Folded Proteins and Intact Protein Complexes by Desorption Electrospray Ionization Mass Spectrometry.

Abstract

Native mass spectrometry (MS) enables the study of intact proteins as well as noncovalent protein-protein and protein-ligand complexes in their biological state. In this work, we present the application of a Waters desorption electrospray ionization (DESI) source with a prototype spray emitter for rapid surface measurements of folded and native protein structures. A comparison of DESI spray solvent shows that adding 50% methanol to 200 mM ammonium acetate solution does not reduce its performance in preserving folded protein structures. Instead, improved signal-to-noise (S/N) ratio is obtained, and less adducted peaks are detected by using this uncommon native MS solvent system. The standard DESI design with an inlet tube allows optimization of sampling temperature conditions to improve desolvation and therefore S/N ratio. Furthermore, tuning the inlet temperature enables the control and study of unfolding behavior of proteins from surface samples. The optimized condition for native DESI has been applied to several selected proteins and protein complexes with the molecular weight ranging from 8.6 to 66.4 kDa. Ions of folded proteins with narrow charge state distribution (CSD), or peaks showing noncovalent-bond-assembled intact protein complexes, are observed in the spectra. Evidence for the structural refolding of denatured proteins and protein complexes sampled with native solvent highlights the need for care when interpreting DESI native MS data, particularly for proteins with stable native structures.