Probing heat and oxidation induced conformational changes of molecular chaperone artemin by excitation-emission fluorescence spectroscopy

Artemin is a potent molecular chaperone, which protects Artemia embryos undergoing encystment against extreme environmental stresses. In the present work, we have examined the structural changes of artemin from A. urmiana upon exposure to oxidant and heat, by using CD measurements as well as excitation-emission fluorescence spectroscopy as a powerful tool for monitoring the conformational transitions and molecular interactions in proteins. We have also provided here the first document on reporting the three dimensional fluorescence spectra of a protein using ANS. Totally, the fluorescence results indicated that the microenvironments of tyrosine and tryptophan residues and the hydrophobic pockets as well as the polypeptide backbone or secondary structure of the chaperone were influenced in responses to heat and H₂O₂ in different degrees. Moreover, the native state of artemin did not induce a considerable exposure of the internal non-polar groups to the solvent. Besides, the excitation-emission spectra of heated artemin by ANS revealed new emission peaks at 430-450 nm when it was excited at 330 nm, which suggests probable exposure of new binding sites for hydrophobic or electrostatic interactions of the protein with ANS. The protein also showed a greater conformational sensitivity to the temperature fluctuations compared to oxidation. Here, we presented some evidence in support of the relation between artemin and its stress dependent activation in vitro and in vivo. This study can expect that the EEM fluorescence spectroscopy could provide a promising tool to study conformational transitions of proteins.