Quantification of Anti-Aggregation Activity of Chaperones: A Test-System Based on Dithiothreitol-Induced Aggregation of Bovine Serum Albumin

The methodology for quantification of the anti-aggregation activity of protein and chemical chaperones has been elaborated. The applicability of this methodology was demonstrated using a test-system based on dithiothreitol-induced aggregation of bovine serum albumin at 45 degrees C as an example. Methods for calculating the initial rate of bovine serum albumin aggregation (v(agg)) have been discussed. The comparison of the dependences of v(agg) on concentrations of intact and cross-linked alpha-crystallin allowed us to make a conclusion that a non-linear character of the dependence of v(agg) on concentration of intact alpha-crystallin was due to the dynamic mobility of the quaternary structure of alpha-crystallin and polydispersity of the alpha-crystallin-target protein complexes. To characterize the anti-aggregation activity of the chemical chaperones (arginine, arginine ethyl ester, arginine amide and proline), the semi-saturation concentration [L](0.5) was used. Among the chemical chaperones studied, arginine ethyl ester and arginine amide reveal the highest anti-aggregation activity ([L](0.5) = 53 and 58 mM, respectively).