Abstract: Histidine Hydrogen Deuterium Exchange Mass Spectrometry (His-HDX MS) determines the HDX rates at the imidazole C2 hydrogen of histidine residues as it’s HDX rate is significantly slower than the others. This property allows the HDX rate to evaluate the state of each histidine residue. We foresee that His-HDX MS has the potential to identify protein ligand interadrawbacks.
Incubation of p38alpha and the inhibitor in the deuterium oxide buffer at 40 to 70 ℃. After the incubation, the deuterated proteins were digested by trypsin, and the resulting digests were analyzed by LC-MS/MS. The obtained mass spectrometry data were analyzed with Byos® (Protein Metrics) HDX workflow, where the extent of deuterium incorporation into His-containing peptides was calculated from the shift of center of m/z.