Interaction of alpha-lactalbumin with mini-alphaA-crystallin

alphaA-Crystallin can function like a molecular chaperone. We have recently shown that residues 71-88 in alphaA-crystallin represent the "chaperone active site" of the protein. A peptide containing the sequence of alphaA-crystallin sequence DFVIFLDVKHFSPEDLTVK (mini alphaA-crystallin) by itself displays the antiaggregation property of alphaA-crystallin. We have prepared a complex of reduced alpha-lactalbumin and mini-alphaA-crystallin and investigated the nature, conformation, and properties of the complex by dynamic light scattering, HPLC analysis, CD spectroscopy, and fluorescence studies. Although mini-alphaA was able to prevent the precipitation of reduced alpha-lactalbumin, large aggregates (50-500 nm) of the complex were formed during the assay. Amino acid composition estimation revealed that alpha-lactalbumin and mini-alphaA-crystallin were present in 1:2 ratio in the aggregates. During our study significant red shift in the Trp fluorescence emission maximum and an increase in Bis-ANS binding to the mini alphaA-crystallin-bound alpha-lacatalbumin were observed. The CD spectra of the complex showed a significant loss of alpha-helical content but the beta-sheet content appeared to be less affected, indicating the molten-globule state of the reduced lactalbumin in the complex. These data show that the active site of alphaA-crystallin by itself can maintain a significantly denatured and unfolded protein in soluble form.