Selective Probing of Acidic Amino Acid Residues for Protein Structural Analysis by Covalent Labeling and Mass Spectrometry

-terminal specific carboxylic acid modification achievable at lower pH. Notably, this approach offers a streamlined conversion of acids to esters, producing molecular nitrogen as the sole byproduct. Validation through tandem mass spectrometry confirms the fragmentation characteristics of labeled peptide samples. Using model peptides and proteins, including bradykinin, neurotensin, angiotensin I, bovine serum albumin (BSA), and the cat allergen Fel d 1 protein complex, we showcase the versatility and applicability of our method for probing reactive acidic residues in structural biology studies. This innovation opens avenues for precise labeling of Glu and Asp residues, providing invaluable insights into protein structure and function, as well as identifying potential therapeutic targets.