Binding of Retinoids to β-Lactoglobulin Isolated by Bioselective Adsorption

Abstract

Binding of the retinoids, all-trans-retinol, all-trans-retinal, all-trans-retinyl acetate, and all-trans-retinoic acid, to beta-lactoglobulin (LG) (96% purity) that had been prepared by bioselective adsorption on N-retinyl-Celite was determined from changes in the fluorescence quenching (332 nm) of the protein tryptophanyl residues. High affinity binding of all of these compounds occurred at pH 7.0, and the apparent dissociation constant ranged from 1.7 to 3.6 x 10(-8) M. Furthermore, a stoichiometry of 1.0 mol.mol-1 of protein was obtained for each case, indicating that all of the sites in the protein preparation were available. When beta-LG in whey protein isolate (57.4% beta-LG) was studied, a stoichiometry of 0.65 to 0.82 mol.mol-1 of protein was obtained, indicating that a large number of the sites already had bound lipid or that the protein had been denatured. As the pH was lowered toward 5.15, the affinity decreased about fourfold, but the stoichiometry of binding was unchanged. Far UV circular dichroism spectra indicated that the secondary structure of the protein was not significantly affected by ligand binding; however, the near UV spectra were changed, indicating that the flexibility of tryptophanyl residues decreased. The latter effect is consistent with the change in fluorescence quenching and suggests that a tryptophan is in the binding site.

Vitamin D3 fortification and quantification in processed dairy products

“…Secondly, vitamin D 3 has been shown to bind to b-lactoglobulin (b-LG), which is a key whey protein. Wang, Allen, and Swaisgood (1997) and Forrest, Yada, and Rousseau (2005) reported the affinity of b-LG for vitamin D 3 . Table 3 shows the percentage retention of vitamin D 3 in cheese over a three-month ripening period.…”

Loss of structural integrity and hydrophobic ligand binding capacity of acetylated and succinylated bovine β-lactoglobulin

“…Extrinsic fluorescence studies were carried out with a conformation-sensitive hydrophobic probe, ANS. In b-lg, hydrophobic ligands, fatty acid and triglycerides binding site have also been established crystallographically inside the hydrophobic calyx (Qin, Creamer, Baker, & Jameson, 1998;Wang et al, 1997;Wu et al, 1999).…”

Maillard glycation of β-lactoglobulin induces conformation changes

“…Crystallographic analysis of the BLG 3-D structure has shown a hydrophobic pocket formed by eight-stranded anti-parallel b-barrel [5,6]. This protein was studied intensively because of its interesting capacity to bind numerous aromatic compounds [7][8][9], retinoids [10][11][12], vitamin D and cholesterol [13], benzopyrene, ellipticine and cisparinaric acid [14,15], vitamin A [16], and fatty acids [17,18]. Although BLG is able to bind a lot of compounds in vitro, its function in milk is still unknown.…”