Temperature-induced folding changes of beta-lactoglobulin in hydro-methanolic solutions

It has been demonstrated using circular dichroism that methanol induces important structural changes of beta-lactoglobulin (BLG). The secondary structure of BLG dissolved in 45% methanol (v/v) at 30 degrees C (dielectric constant epsilon approximately 50) is predominantly alpha-helical and unable to complex retinol any more. However, when the dielectric constant of such a medium is raised back to epsilon approximately 70 by decreasing the temperature, both the refolding of BLG into a beta-structure and the formation of the retinol/BLG complex are observed. On the other hand, in the case of BLG solution in 30% methanol, the decrease of the dielectric constant from epsilon approximately 69 to epsilon approximately 53 by heating from 20 degrees C to 60 degrees C leads to the transition from a predominantly beta-structure into a predominantly random one and the dissociation of the retinol/BLG complex. The reversibility of the conformation changes by cooling was demonstrated by circular dichroism curves and tryptophan fluorescence. The retinol fluorescence intensity could not be completely recovered.