Identification of glycated and acetylated lysine residues in human α2-antiplasmin

Abstract Background The post-translational protein modification via lysine residues can significantly alter its function. α2-antiplasmin, a key inhibitor of fibrinolysis, contains 19 lysine residues. Aim We sought to identify sites of glycation and acetylation in human α2-antiplasmin and test whether the competition might occur on the lysine residues of α2-antiplasmin. Methods We analyzed human α2-antiplasmin (1) untreated; (2) incubated with increasing concentrations of β- d -glucose (0, 5, 10, 50 mM); (3) incubated with 1.6 mM acetylsalicylic acid (ASA) and (4) incubated with 1.6 mM ASA and 50 mM β- d -glucose, using the ultraperformance liquid chromatography system coupled to mass spectrometer. Results Eleven glycation sites and 10 acetylation sites were found in α2-antiplasmin. Incubation with β- d -glucose was associated with glycation of 4 (K-418, K-427, K-434, K-441) out of 6 lysine residues, known to be important for mediating the interaction with plasmin. Glycation and acetylation overlapped at 9 sites in samples incubated with β- d -glucose or ASA. Incubation with concomitant ASA and β- d -glucose was associated with the decreased acetylation at all sites overlapping with glycation sites. At K-182 and K-448, decreased acetylation was associated with increased glycation when compared with α2-antiplasmin incubated with 50 mM β- d -glucose alone. Although K-24 located in the proximity of the α2-antiplasmin cleavage site, was found to be only acetylated, incubation with ASA and 50 mM β- d -glucose was associated the absence of acetylation at that site. Conclusion Human α2-antiplasmin is glycated and acetylated at several sites, with the possible competition between acetylation and glycation at K-182 and K-448. Our finding suggests possibly relevant alterations to α2-antiplasmin function at high glycemia and during aspirin use.