Hydrogen/Deuterium Exchange Reaction Rate of Cytochrome c Determined by Droplet Collision Atmospheric Pressure Infrared Laser Ablation Mass Spectrometry.

The hydrogen/deuterium (H/D) exchange rate is an optimal measure for studying the structures and dynamics of hydrogen bonding systems, as it reflects the molecular contact environment and the strength of the hydrogen bonds. A method for rapid measurement of the H/D exchange reaction rates is required to examine the intermolecular environments of molecules in solutions. We developed a droplet collision atmospheric pressure infrared laser ablation mass spectrometry technique for this purpose. We obtained the H/D exchange reaction rate of cytochrome c in a methanol/H ₂ O·D ₂ O solution. We revealed that the first hydration shell of the cytochrome c molecule hinders the penetration of D ₂ O to the surface of the molecule from the rates, which provides a novel method to investigate solution structures by a mass-spectrometric method. The droplet-collision mass spectrometry method developed in the present study can be extended to research on the molecular interactions in solutions, such as the mutual interactions of protein molecules, which are of importance in living cells.