Molecular characterization of alpha-lactalbumin folding variants that induce apoptosis in tumor cells

This study characterized a protein complex in human milk that induces apopt osis in tumor cells but spares healthy cells. The active fraction was purif ied from casein by anion exchange chromatography. Unlike other casein compo nents the active fraction was retained by the ion exchanger and eluted afte r a high salt gradient. The active fraction showed N-terminal amino acid se quence identity with human milk alpha-lactalbumin and mass spectrometry rul ed out post-translational modifications. Size exclusion chromatography reso lved monomers and oligomers of alpha-lactalbumin that were characterized us ing UV absorbance, fluorescence, and circular dichroism spectroscopy. The h igh molecular weight oligomers were kinetically stable against dissociation into monomers and were found to have an essentially retained secondary str ucture but a less well organized tertiary structure. Comparison with native monomeric and molten globule alpha-lactalbumin showed that the active frac tion contains oligomers of alpha-lactalbumin that have undergone a conforma tional switch toward a molten globule-like state. Oligomerization appears t o conserve alpha-lactalbumin in a state with molten globule-like properties at physiological conditions. The results suggest differences in biological properties between folding variants of alpha-lactalbumin.