Conversion of Human α-lactalbumin to an Apo-like State in the Complexes with Basic Poly-Amino Acids:  Toward Understanding of the Molecular Mechanism of Antitumor Action of HAMLET

It was recently shown that α-lactalbumin associated with oleic acid (HAMLET) interacts with core histones thereby triggering apoptosis of tumor cells (J. Biol. Chem. 2003, 278, 42131). In previous work, we revealed that monomeric α-lactalbumin in the absence of fatty acids can also interact with histones and, moreover, with basic poly-amino acids (poly-Lys and poly-Arg) that represent simple models of histone proteins (Biochemistry 2004, 43, 5575). Association of α-lactalbumin with histone or poly-Lys(Arg) essentially changes its properties. In the present work, the character of the changes in structural properties and conformational stability of α-lactalbumin in the complex with poly-Lys(Arg) has been studied in detail by steady-state fluorescence, circular dichroism, and differential scanning calorimetry. Complex formation strongly depends on ionic strength, confirming its electrostatic nature. Experiments with the poly-amino acids of various molecular masses demonstrated a direct proportionality between the number of α-lactalbumin molecules bound per poly-Lys(Arg) and the surface area of the poly-amino acid random coil. The binding of the poly-amino acids to Ca2+-saturated human α-lactalbumin decreases its thermal stability down to the level of its free apo-form and decreases Ca2+-affinity by 4 orders of magnitude. The conformational state of α-lactalbumin in a complex with poly-Lys(Arg), named α-LActalbumin Modified by Poly-Amino acid (LAMPA), differs from all other α-lactalbumin states characterized to date, representing an apo-like (molten globule-like) state with substantially decreased affinity for calcium ion. The requirement for efficient conversion of α-lactalbumin to the LAMPA state is a poly-Lys(Arg) chain consisting of several tens of amino acid residues. Keywords: α-lactalbumin • histones • poly-amino acids • electrostatics • thermal stability • calcium-binding