Rapid Assessment of Pepsin Column Activity for Reliable HDX-MS Studies.
Clint VorauerMichael S WrigleyJuan P Rincon PabonMichael J WatsonCharlie C MundorffDavid D WeisMiklos M GuttmanPublished in: Journal of the American Society for Mass Spectrometry (2021)
Hydrogen/deuterium exchange with mass spectrometry (HDX-MS) is a widely used technique to probe protein structural dynamics, track conformational changes, and map protein-protein interactions. Most HDX-MS studies employ a bottom-up approach utilizing the acid active protease pepsin to digest the protein of interest, often utilizing immobilized protease in a column format. The extent of proteolytic cleavage will greatly influence data quality and presents a major source of variation in HDX-MS studies. Here, we present a simple cocktail of commonly available peptides that are substrates of pepsin and can serve as a rapid check of pepsin column activity. The peptide-based assay requires no system modifications and provides an immediate readout to check and benchmark pepsin activity across different HDX-MS platforms.
Keyphrases
- mass spectrometry
- liquid chromatography
- ms ms
- multiple sclerosis
- capillary electrophoresis
- gas chromatography
- high performance liquid chromatography
- tandem mass spectrometry
- high resolution
- case control
- amino acid
- solid phase extraction
- molecular dynamics simulations
- quantum dots
- electronic health record
- binding protein
- high throughput
- dna binding
- machine learning
- loop mediated isothermal amplification