Localization of Potential Transglutaminase Cross-Linking Sites in Bovine Caseins

Transglutaminase acyl donor sites in the four individual bovine caseins were determined. Purified caseins labeled with a radioactive site-specific probe in a guinea pig liver transglutaminase-catalyzed reaction were subjected to enzymatic digestion followed by reversed-phase HPLC separation of the resulting peptides. By a combination of sequence and mass spectrometric analysis of the major radioactive fractions, the following TG-reactive glutamine residues were found: α s1 -casein, Gln13, Gln108, Gln130, and Gln140; α s2 -casein, Gln79, Gln169, Gln185, and Gln187; β-casein, Gln54, Gln56, Gln72, Gln79, and Gln182; κ-casein, Gln29, Gln45, Gln114, and Gln163. In addition, an intra- or intermolecular cross-link between Gln45 and each of two adjacent lysine residues, Lys21 and Lys24 in monomeric κ-casein, was identified.