Mass spectrometric analysis of asparagine deamidation and aspartate isomerization in polypeptides.

One of the most frequent modifications in proteins and peptides is the deamidation of asparagine, a spontaneous non-enzymatic reaction leading to a mixture of L,D-succinimidyl, L,D-aspartyl, and L,D-isoaspartyl forms, with L-isoaspartyl dominating. Spontaneous isomerization of L-Asp yields the same products. In vivo, these unusual forms of aspartate are repaired by the protein L-isoaspartyl O-methyltransferase enzyme, with the balance between isomerization and repair affecting the organism physiology. Mass spectrometric analysis of this balance involves isomer separation, iso-Asp/Asp quantification, and iso-Asp site identification. This review highlights the issues associated with these steps and discusses the prospects of high-throughput iso-Asp analysis.