Binding of vitamin A with milk α- and β-caseins

The binding sites of retinol and retinoic acid with milk alpha- and beta-caseins were determined, using constant protein concentration and various retinoid contents. FTIR, UV-visible and fluorescence spectroscopic methods as well as molecular modelling were used to analyse retinol and retinoic acid binding sites, the binding constant and the effect of retinoid complexation on the stability and conformation of caseins. Structural analysis showed that retinoids bind caseins via both hydrophilic and hydrophobic contacts with overall binding constants of Kretinol-alpha-caseins = 1.21 (+/- 0.4) x 10(5) M-1 and Kretinol-beta-caseins = 1.11 (+/- 0.5) x 10(5) M-1 and K-retinoic (acid-alpha-caseins) = 6.2 (+/- 0,6) x 10(4) M-1 and K-retinoic (acid-beta-caseins) = 6.3 (+/- 0.6) x 10(4) M-1. The number of bound retinol molecules per protein (n) was 1.5 (+/- 0.1) for alpha-casein and 1.0 (+/- 0.1) for beta-casein, while 1 molecule of retinoic acid was bound in the alpha- and beta-casein complexes. Molecular modelling showed different binding sites for retinol and retinoic acid on alpha- and beta-caseins with more stable complexes formed with alpha-casein. Retinoid-casein complexation induced minor alterations of protein conformation. Caseins might act as carriers for transportation of retinoids to target molecules. (C) 2012 Elsevier Ltd. All rights reserved.