Thermal aggregation of globulin from an indigenous Chinese legume, Phaseolus angularis (red bean)

The thermal aggregation behavior of red bean (Phaseolus angularis) globulin (RBG) was studied at ≈1% (w/v) protein concentration in 0.01 M phosphate buffer, pH 7.4. The percentage of protein precipitated was affected by heating temperature, heating time and salt concentration. The influences of several salts of the chaotropic series and protein structure-modifying agents on thermal coagulation of RBG were also investigated. The effects of chaotropic salts did not follow the lyotropic series of anions. Sodium dodecyl sulfate caused a more pronounced reduction in heat-induced aggregation of RBG than did dithiothreitol, while N-ethylmaleimide did not affect aggregation until after a long heating period. Differential scanning calorimetric (DSC) data showed that heat aggregation of RBG was preceded by thermal denaturation. SDS-PAGE showed that heating led to the disappearance of some protein bands, and the basic polypeptide of 11S globulin (legumin) was not found in the buffer-soluble aggregates. Heating caused increases of surface hydrophobicity, again suggesting protein unfolding prior to aggregate formation. The buffer-insoluble aggregates did not show any DSC response, indicating extensive denaturation, and had a lower surface hydrophobicity and higher disulfide content than the buffer-soluble aggregates. The data suggest that electrostatic and hydrophobic interactions may play an important role in thermal aggregation of RBG, with disulfide bonds playing a limited role.