pH-Dependent Aggregation and Disaggregation of Native β-Lactoglobulin in Low Salt

Abstract

The aggregation of β-lactoglobulin (BLG) near its isoelectric point was studied as a function of ionic strength and pH. We compared the behavior of native BLG with those of its two isoforms, BLG-A and BLG-B, and with that of a protein with a very similar pI, bovine serum albumin (BSA). Rates of aggregation were obtained through a highly precise and convenient pH/turbidimetric titration that measures transmittance to ±0.05 %T. A comparison of BLG and BSA suggests that the difference between pHmax (the pH of the maximum aggregation rate) and pI is systematically related to the nature of protein charge asymmetry, as further supported by the effect of localized charge density on the dramatically different aggregation rates of the two BLG isoforms. Kinetic measurements including very short time periods show well-differentiated first and second steps. BLG was analyzed by light scattering under conditions corresponding to maxima in the first and second steps. Dynamic light scattering (DLS) was used to monitor the kinetics, and static light scattering (SLS) was used to evaluate the aggregate structure fractal dimensions at different quench points. The rate of the first step is relatively symmetrical around pHmax and is attributed to the local charges within the negative domain of the free protein. In contrast, the remarkably linear pH dependence of the second step is related to the uniform reduction in global protein charge with increasing pH below pI, accompanied by an attractive force due to surface charge fluctuations.

Control of thermal fabrication and size of β-lactoglobulin-based microgels and their potential applications

“…Above 30 mmol kg À1 ionic strength the temperature of maximum turbidity development increased. This reduction in aggregation with increased ionic strength agrees with earlier reports on the reduced interactivity between Blg molecules in the presence of ions [40]. Visible aggregates were observed in microgel systems with added ionic strength in excess of 30 mmol kg À1 , which contributed to the observed reduction in peak heights and the change in the aggregation profiles under these conditions.…”

Heteroprotein complex coacervation: A generic process

“…α-LA also presents a very specific calcium binding site (Ka = 10 8 M -1 ) formed by the residues 79 to 88 [17]. a more acidic and hydrophobic character than variant B and this explains their differential aggregation properties found near β-LG isoelectric point [21]. β-LG secondary structure is composed of about 10-15% α-helix, 43% β-sheet and 47% unordered structure including β-turn [22].…”

“…ACCEPTED MANUSCRIPT 21 and caseins, either CN-micelle or purified -casein (-CN) has been the focus of some research groups.…”

Current ways to modify the structure of whey proteins for specific functionalities—a review

“…The remaining free thiol group is buried within the protein structure on the β-strand H. The disulfide bonds contribute to the reversible denaturation of β-LG (Kitabatake et al 2001), whereas the free thiol stabilizes the native protein structure (Jayat et al 2004). At room temperature, β-LG exhibits pH-dependent reversible self-association behavior (Yan et al 2013). At neutral pH (5.5-7.5), a stable noncovalent dimer is formed, whereas around its pI (5.2), β-LG exists as octamer.…”