X-ray structural evidence for a local helix-loop transition in alpha-lactalbumin

The three-dimensional structure of human alpha-lactalbumin for two crystal forms has been determined by x-ray analysis. One crystal (the form LT) was obtained at pH 4.2 and room temperature, while the other crystal (the form HT) was grown at pH 6.5 and 37 degrees C. The backbone structure for Lys1-Ile95 residues is almost conserved between the two structures as indicated by the root mean square difference of 0.30 A for the superposition of equivalent C alpha atoms. The calcium ion is surrounded by seven oxygen atoms of three carboxyl groups, two carbonyl groups, and two water molecules, which form a distorted pentagonal bipyramid in both structures. A large difference in polypeptide folding is found in the region of Leu96-Leu123 residues. Especially in the region of Trp104-Cys111 residues, a distorted alpha-helix is observed in the form HT while a loop structure is formed in the other crystal. The fact that the crystals of both forms appeared in the same batch at pH 6.5 and room temperature indicates that the human alpha-lactalbumin structure is highly fluctuated in solution and the folding and unfolding of the alpha-helix of Trp104-Cys111 residues are in equilibrium. Since the crystal of the form HT exclusively appeared around the physiological temperature, the structure of this form can be considered as the native structure. The partially unfolded structure in the form LT indicates that the local denaturation occurs even at room temperature.