Antibiotic doxorubicin and its derivative bind milk β-lactoglobulin.

β-Lactoglobulin (β-LG) is a member of lipocalin superfamily of transporters for small hydrophobic molecules such as doxorubicin and its derivatives. We located the binding sites of doxorubicin (DOX) and N-(trifluoroacetyl) doxorubicin (FDOX) with β-lactoglobulin in aqueous solution at physiological conditions, using FTIR, CD and fluorescence spectroscopic methods as well as molecular modeling. Structural analysis showed that DOX and FDOX bind β-LG via both hydrophilic and hydrophobic contacts with overall binding constants of K(DOX-)(β)(-LG)=1.0 (± 0.4)× 10(4)M(-1) and K(FDOX-)(β)(-LG)=2.5 (± 0.5)× 10(4)M(-1) and the number of drug molecules bound per protein (n) 1.2 for DOX and 0.6 for FDOX. Molecular modeling showed the participation of several amino acids in the drug-protein complexes with the free binding energy of -8.12 kcal/mol for DOX-β-LG and -7.74 kcal/mol for FDOX-β-LG complexes. DOX and FDOX do not share similar binding sites with β-LG. Protein conformation showed minor alterations with reduction of β-sheet from 58% (free protein) to 57-51% in the drug-β-LG complexes. β-LG can transport doxorubicin and its derivative in vitro.