Localizing flexible regions in proteins using hydrogen-deuterium exchange mass spectrometry

Hydrogen-deuterium exchange (HDX) can provide invaluable structural information for proteins. The incorporation of deuterium into a protein's backbone amide is readily monitored by mass spectrometry (MS). Assuming that the molecular weight of the protein is not a limiting factor of the MS, HDXMS can be performed on intact proteins; however, digesting the protein prior to MS allows one to assign HDX information to specific peptides within the protein. Here, we describe HDXMS data collection and analysis to identify regions based on their degree of protection in the pharmaceutical protein glucocerebrosidase (GCase).