Characterization of intermediates formed during heat-induced aggregation of β-lactoglobulin AB at neutral pH

Heat-induced aggregation of beta-lactoglobulin AB (17 mg mL(-1) in 0.005 M NaCl; pH 7.0; 78.5 degrees C) was studied using size-exclusion chromatography in combination with multi-angle laser light scattering and electrophoretic techniques. By choosing the chromatographic condition such that the native beta-lactoglobulin was partially associated, the formation of irreversibly altered monomers and non-native dimers could be monitored. Heat-induced aggregation of beta-lactoglobulin occurred via many intermediates, and mainly via disulphide bonding and to a much lesser extent via non-covalent interactions. In the early stages of the aggregation of beta-lactoglobulin non-native dimers and oligomers were formed, presumably mainly via thiol-disulphide exchange and to a lesser extent via thiol-thiol oxidation and non-covalent interactions. In later stages aggregates with a continuous distribution of molar masses from 10(5) to 2 x 10(6) Da were formed, presumably via the incorporation of monomers and small aggregates into larger aggregates mainly through disulphide bonding. (C) 2000 Elsevier Science Ltd. All rights reserved.