Rapid Analysis of Protein Structure and Dynamics by Hydrogen/Deuterium Exchange Mass Spectrometry

摘要: An automated approach for the rapid analysis of protein structure has been developed and used to study acid-induced conformational changes in human growth hormone. The labeling involves hydrogen/deuterium exchange (H/D-Ex) backbone amide hydrogens with sensitive detection by mass spectrometry (MS). Briefly, is incubated defined intervals a deuterated environment. After quenching reaction, partially enzymatically digested resulting peptide fragments are analyzed liquid chromatography (LC-MS). deuterium buildup curve measured each fragment yields an average rate that reflects environment intact protein. Additional analyses allow mapping free energy folding on localized segments along sequence affording unique dynamic structural information. While H/D-Ex coupled MS recognized as powerful technique studying protein–ligand interactions, it remained labor-intensive task. improvements methodology described here include solid phase proteolysis, handling sample preparation, integrated data reduction software together improve coverage resolution, while achieving throughput nearly 10-fold higher than commonly manual methods.