Structural differences between bovine A1 and A2 β-casein alter micelle self-assembly and influence molecular chaperone activity

Abstract

Within each milk protein there are many individual protein variants and marked alterations to milk functionality can occur depending on the genetic variants of each protein present. Bovine A(1) and A(2) β-casein (β-CN) are 2 variants that contribute to differences in the gelation performance of milk. The A(1) and A(2) β-CN variants differ by a single AA, the substitution of histidine for proline at position 67. β-Casein not only participates in formation of the casein micelle but also forms an oligomeric micelle itself and functions as a molecular chaperone to prevent the aggregation of a wide range of proteins, including the other caseins. Micelle assembly of A(1) and A(2) β-CN was investigated using dynamic light scattering and small-angle X-ray scattering, whereas protein functionality was assessed using fluorescence techniques and molecular chaperone assays. The A(2) β-CN variant formed smaller micelles than A(1) β-CN, with the monomer-micelle equilibrium of A(2) β-CN being shifted toward the monomer. This shift most likely arose from structural differences between the 2 β-CN variants associated with the adoption of greater polyproline-II helix in A(2) β-CN and most likely led to enhanced chaperone activity of A(2) β-CN compared with A(1) β-CN. The difference in micelle assembly, and hence chaperone activity, may provide explain differences in the functionality of homozygous A(1) and A(2) milk. The results of this study highlight that substitution of even a single AA can significantly alter the properties of an intrinsically unstructured protein such as β-CN and, in this case, may have an effect on the functionality of milk.

Casein polymorphism heterogeneity influences casein micelle size in milk of individual cows

“…Our recent work on purified A 1 and A 2 structural selfassembly behavior has also indicated that the genetic diversity of β-CN might influence the internal structure of casein micelle assembly and possibly monomer-micelle equilibrium (Raynes et al, 2015). Nonhomogeneity of β-CN could limit the symmetric assembly and growth of casein micelles.…”

The molecular chaperone β-casein prevents amorphous and fibrillar aggregation of α-lactalbumin by stabilisation of dynamic disorder

“…Herein, we investigated the effects of the molecular chaperone β-CN on both amorphous and amyloid fibrillar aggregation of α-LA as representative examples of protein unfolding and its mitigation. In a broader context, the study also has relevance to the dairy industry as the chaperone action of β-CN is important in stabilising other milk proteins such as α-LA, for example under ultra-high temperature and pasteurisation processing [22,39].…”

Investigation of Age Gelation in UHT Milk

“…β-CN A1 and A2 differ by one amino acid at position 67, His to Pro, respectively. This substitution produces different protease cleavage products upon simulated gastrointestinal digestion [19], and we showed that A2 β-CN forms into a smaller self-associating micelle and functions more efficiently as a molecular chaperone to inhibit protein aggregation [20]. β-Lg A and B differ at position 64, Asp to Gly, and 118, Val to Ala, respectively.…”