Organic Solvents for Enhanced Proteolysis of Stable Proteins for Hydrogen-Deuterium Exchange Mass Spectrometry.

ABSTRACT: Protein digestion is a key challenge in mass spectrometry (MS)-based structural proteomics. Although using hydrogen-deuterium exchange kinetics with MS (HDX-MS) to interrogate the high-order structure of proteins is now established, it can be challenging for β-barrel proteins, which are important in cellular transport. These proteins contain a continuous chain of H-bonds that impart stability, causing difficulty in digestion for bottom-up measurements. To overcome this impediment, we tested organic solvents as denaturants during on-line pepsin digestion of soluble β-barrel proteins. We selected green fluorescent protein (GFP), siderocalin (Scn), and retinol-binding protein 4 (RBP4) as model proteins and screened six different polar-aprotic and polar-protic solvent combinations to disrupt t

SUBMITTER: Guo C 

PROVIDER: S-EPMC7485609 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature