Involvement of lysine residues of goat serum albumin in high-affinity binding of bilirubin.

Four maleylated derivatives of goat serum albumin having percent modification as 40%, 46%, 84% and 98% were prepared using varying molar ratio of maleic anhydride over protein. These preparations were found to be pure, both with respect to size and charged as judged by gel filtration and polyacrylamide gel electrophoresis. Maleylation caused significant change in protein conformation as revealed by the change in Stokes radius and frictional ratio of serum albumin, from 3.46 nm and 1.28 to 4.96 nm and 1.79, respectively, upon 98% modification. Immunodiffusion results of native and modified albumins with anti-goat serum albumin antiserum also suggested significant conformational changes in serum albumin upon maleylation. About 88% reduction in bilirubin binding was observed after modification of 98% amino groups of serum albumin as studied by visible difference spectroscopy at pH 8.0, and at 0.15 ionic strength. Increase in ionic strength to 1.0 did not lead to any significant reversal in bilirubin binding. These results prove the involvement of lysine residues in bilirubin-albumin interaction.