Subzero Celsius separations in three-zone temperature controlled hydrogen deuterium exchange mass spectrometry

摘要

Highlights?HDX MS subzero separations via an addition to widely available hardware.?There were no major changes to chromatographic efficiency at ?10°C and ?20°C.?Significant improvements in deuterium recovery were achieved at subzero temperatures.AbstractHydrogen deuterium exchange mass spectrometry (HDX MS) reports on the conformational landscape of proteins by monitoring the exchange between backbone amide hydrogen atoms and deuterium in the solvent. To maintain the label for analysis, quench conditions of low temperature and pH are required during the chromatography step performed after protease digestion but before mass spectrometry. Separation at 0°C is often chosen as this is the temperature where the most deuterium can be recovered without freezing of the typical water and acetonitrile mobile phases. Several recent reports of separations at subzero Celsius emphasize the promise for retaining more deuterium and using a much longer chromatographic gradient or direct infusion time. Here we present