Thermal Aggregation of β-Lactoglobulin: Effect of pH, Ionic Environment, and Thiol Reagent

Heat-induced β-lactoglobulin (1.2 mg/ml) aggregation in NaCl, CaCl₂, and thiol-blocking agent (N-ethylmaleimide) solutions was determined by measuring dynamic turbidity changes at temperatures ranging from 25 to 96°C. β-Lactoglobulin in distilled water exhibited a single transition (≥76°C) in protein-protein interaction (aggregation). An increase in pH from 5.50 to 6.50 suppressed the transition, whereas addition of NaCl (.02 to 1.0 M) and CaCl₂ (.005 to .2 M) promoted the transition. N-Ethylmaleimide (4.0 to 10.0 mM) decreased the transition temperature and, in the absence of salts, induced a second transition. However, the combination of N-ethylmaleimide and NaCl or CaCl₂ produced a single, large transition peak. Results indicate that β-lactoglobulin aggregation is most sensitive to low pH, greatly depends on the type and concentration of specific salts, and involves electrostatic and possibly hydrophobic forces and sulfhydryl reactions.