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.
Sreelakshmi
Y
Y
Department of Ophthalmology, University of Missouri, Columbia 65212, USA.
Sharma
K K
KK
D000817:Anilino Naphthalenesulfonates / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D011232:Chemical Precipitation; D002942:Circular Dichroism; D003459:Crystallins / Q000737:chemistry*; D005453:Fluorescence; D006863:Hydrogen-Ion Concentration; D007768:Lactalbumin / Q000737:chemistry*; D018832:Molecular Chaperones / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation / Q000187:drug effects; D014364:Tryptophan / Q000737:chemistry
D000817:Anilino Naphthalenesulfonates; D003459:Crystallins; D018832:Molecular Chaperones; C035763:5,5'-bis(8-(phenylamino)-1-naphthalenesulfonate); D014364:Tryptophan; D007768:Lactalbumin