Effect of heat treatment on the conformation and aggregation of beta-lactoglobulin A, B, and C

Solutions of bovine beta-lactoglobulin (beta-Lg) A-C were heated at tempera tures between 50 and 90 degrees C for 12.5 min at pH 6.7 or 7.4, and the pr oducts were analyzed by alkaline, sodium dodecyl sulfate (SDS), and two-dim ensional (2D) (alkaline and then SDS) polyacrylamide gel electrophoresis (P AGE). Results from the pH 6.7 samples that were similar to 70% denatured sh owed that the proportion of beta-Lg that was in very large aggregates was b eta-Lg C > beta-Lg B > beta-Lg A. 2D PAGE showed that there were a large nu mber of unexpected intermediate products, especially from beta-Lg A. These and other results, including the dissociation of disulfide-bonded dimers fr om trimers and tetramers by SDS, indicate that (1) beta-Lg dimers could be important intermediates in the further aggregation of beta-Lg, (2) hydropho bically driven associations occur within the aggregates, (3) the mechanism of beta-Lg aggregation is not simple, and (4) differences in variant protei n behavior are explainable in terms of net negative charge and specific ami no acid substitutions.