Pressure-induced unfolding and aggregation of the proteins in whey protein concentrate solutions

Abstract

Whey protein concentrate solutions (12% w/v, pH 6.65 ± 0.05) were pressure treated at 800 MPa for 20-120 min and then examined using size exclusion chromatography (SEC), small deformation rheology, transmission electron microscopy, and various types of one-dimensional (1D) and two-dimensional (2D) polyacrylamide gel electrophoresis (PAGE). The pressure-treated samples showed a time-dependent loss of native whey proteins by SEC and 1D PAGE and a corresponding increase in non-native proteins and protein aggregates of different sizes. These aggregates altered the viscosity and opacity of the samples and were shown to be cross-linked by intermolecular disulfide bonds and by noncovalent interactions using 1D PAGE [alkaline (or native), sodium dodecyl sulfate (SDS), and SDS of reduced samples (SDS R)] and 2D PAGE (native:SDS and SDS:SDSR). The sensitivity of the major whey proteins to pressure was in the order β-lactoglobulin B (β-LG B) > β-LG A > bovine serum albumin (BSA) > α-lactalbumin (α-LA), and the large internal hydrophobic cavity of β-LG may have been partially responsible for its sensitivity to high-pressure treatments. It seemed likely that, at 800 MPa, the formation of a β-LG disulfide-bonded network preceded the formation of disulfide bonds between α-LA or BSA and β-LG to form multiprotein aggregates, possibly because the disulfide bonds of α-LA and BSA are less exposed than those of β-LG either during or after pressure treatment. It may be possible that intermolecular disulfide bond formation occurred at high pressure and that hydrophobic association became important after the high-pressure treatment. © 2005 American Chemical Society.