Investigating Protein-Nucleic Acid Binding Interactions with Diethylpyrocarbonate Covalent Labeling–Mass Spectrometry

• Published in: Journal of the American Society for Mass Spectrometry 2024-10, Vol.35 (10), p.2272-2275
• Main Authors: Kirsch, Zachary J., Ashby, Jonathan, Vachet, Richard W.
• Format: Article
• Language: English
• ISSN: 1044-0305; 1879-1123; 1879-1123
• DOI: 10.1021/jasms.4c00285

Nucleic acids are important biomolecules that facilitate numerous cellular functions and have in recent years become promising candidates for treating disease. Consequently, there is a need for methods to characterize protein interactions with these molecules. Here, we demonstrate that diethylpyrocarbonate (DEPC) covalent labeling–mass spectrometry (CL-MS) can provide structural information for protein-nucleic acid binding by characterizing the binding sites of two DNA aptamers specific to thrombin. Reductions in thrombin labeling are observed at the pair’s binding interfaces. Furthermore, we find that binding of the aptamers causes changes in labeling at residues in the thrombin active site and known exosites for each aptamer, showcasing the sensitivity of DEPC CL-MS to significant allosteric changes.