β-Casein will chaperone β-lactoglobulin during nanofibril assembly, but prefers familiar company at high concentrations

Bovine β-casein behaves as a chaperone at neutral pH, i.e., it inhibits aggregation of unfolded proteins, but chaperone effects have not been investigated at acidic pH. We examined chaperone effects during heating at pH 2 and 80 °C for mixtures of 1% (w/v) β-lactoglobulin (β-Lg) and β-Lg:β-casein molar ratios of 1:0.0625 to 1:1. These conditions hydrolyse β-Lg into peptides that self-assemble into amyloid-like nanofibrils. Hydrolysis rates in mixed solutions were unchanged from single-protein controls, according to sodium dodecyl sulphate polyacrylamide gel electrophoresis. At ratios ≥1:0.125, β-casein slowed fibril growth in a concentration-independent way, according to Thioflavin T fluorescence. We observed twisted irregular fibrils coexisting with long semiflexible fibrils when β-casein was heated with β-Lg, using transmission electron microscopy. We hypothesise that β-casein monomers and peptides can interact with growing β-Lg nanofibrils or assemble into micelles, with the latter pathway predominating at high β-casein concentrations.