Limited Lactosylation of Beta-Lactoglobulin from Cow’s Milk Exerts Strong Influence on Antigenicity and Degranulation of Mast Cells

• Published in: Nutrients 2021-06, Vol.13 (6), p.2041
• Main Authors: Bosman, Gerlof P, Oliveira, Sergio, Simons, Peter J, Sastre Torano, Javier, Somsen, Govert W, Knippels, Leon M. J, Haselberg, Rob, Pieters, Roland J, Garssen, Johan, Knipping, Karen
• Format: Article
• Language: English
• Subjects: Allergenicity; Allergens; Antibodies; Antigenicity; beta-lactoglobulin; Binding; Capillary electrophoresis; Cow's milk; cow’s milk allergy; Degranulation; Electrophoresis; Enzyme-linked immunosorbent assay; Food; Food allergies; Heat; High temperature; Immunoglobulin E; Immunoglobulin G; Laboratories; Lactoglobulin; Lactose; lactosylation; Maillard reaction; Mass spectrometry; Mass spectroscopy; mast cell degranulation; Mast cells; Milk; N-Terminus; Pharmaceutical industry; Proteins; Scientific imaging; Sugar; Water activity; Whey; β-Lactoglobulin
• ISSN: 2072-6643; 2072-6643
• DOI: 10.3390/nu13062041

Background: beta-lactoglobulin (BLG) is one of the major cow’s milk proteins and the most abundant allergen in whey. Heating is a common technologic treatment applied during milk transformational processes. Maillardation of BLG in the presence of reducing sugars and elevated temperatures may influence its antigenicity and allergenicity. Primary objective: to analyze and identify lactosylation sites by capillary electrophoresis mass spectrometry (CE-MS). Secondary objective: to assess the effect of lactosylated BLG on antigenicity and degranulation of mast cells. Methods: BLG was lactosylated at pH 7, a water activity (aw) of 0.43, and a temperature of 65 °C using a molar ratio BLG:lactose of 1:1 by incubating for 0, 3, 8, 16 or 24 h. For the determination of the effect on antibody-binding capacity of lactosylated BLG, an ELISA was performed. For the assessment of degranulation of the cell-line RBL-hεIa-2B12 transfected with the human α-chain, Fcε receptor type 1 (FcεRI) was used. Results: BLG showed saturated lactosylation between 8 and 16 incubation hours in our experimental setup. Initial stage lactosylation sites L1 (N-terminus)—K47, K60, K75, K77, K91, K138 and K141—have been identified using CE-MS. Lactosylated BLG showed a significant reduction of both the IgG binding (p = 0.0001) as well as degranulation of anti-BLG IgE-sensitized RBL-hεIa-2B12 cells (p < 0.0001). Conclusions and clinical relevance: this study shows that lactosylation of BLG decreases both the antigenicity and degranulation of mast cells and can therefore be a promising approach for reducing allergenicity of cow’s milk allergens provided that the process is well-controlled.