Formation of isoaspartyl peptide bonds (isoAsp) is one of the most common forms of non-enzymatic degradation of peptides and proteins under mild conditions. IsoAsp arises when certain Asn–Xaa and Asp–Xaa sites undergo a spontaneous intramolecular rearrangement to form a succinimide which subsequently hydrolyzes to generate a mixture of isoAsp–Xaa and Asp–Xaa linkages in a ratio of ∼2:1. This pathway is responsible for the much greater susceptibility of asparagine, compared with glutamine, to deamidation at neutral and alkaline pH. Rearrangement occurs most readily at Asn–Gly, Asn–Ser, and Asp–Gly sequences where the local polypeptide chain flexibility is high. Formation of isoAsp can decrease the biological activity of a protein pharmaceutical, alter its susceptibility to proteolytic degradation, and elicit autoimmunity. The enzyme protein l-isoaspartyl methyltransferase can be used to measure isoAsp sites in the low pmol range with or without the use of radioisotopes.