DAB389IL-2 with the trade name denileukin diftitox and ontak is the first recombinant immunotoxinthat produced using genetic engineering . In this commercial fusion protein binding domain ofdiphtheria toxin (DT) replaced by the amino acid sequence of human interleukin-2 (IL-2) [2,3]. The aimof this work is high level expression, refolding and characterization of this diphtheria fusion toxin. Forthis, expressed DAB389IL-2 was refolded and purified in thepresence of two anti-aggregators, sorbitol andL-arginine. Refolding
of protein have been investigated using native-PAGE, fluorescence and circulardichroism (CD) spectroscopy. Finally the proper functioning was determined by nuclease activityassay.The results revealed that the fluorescence intensity of protein decreased in the presence of both antiaggregators.Far-UV CD spectra indicated that the protein has some conformational alterations uponinteracting with these anti-aggregators in comparison with the unfolded structure. In fact, in the presenceof two refolding buffer the intensity of far-UV CD signals have been increased at the regions of 208 and222 nm. Moreover, native-gel electrophoresis data demonstrated that the refolded configuration of proteinhas been formed in the both refolding buffers. Finally we have found that the refolded proteins haveproper activity as indicated by nuclease activity. Therefore, addition of sorbitol and L-arginine perhapsincrease the viscosity and surface tension of solvent which leads to increase the strength of hydrogenbonds and then stabilizes the refolded structure.Results of this research could give more insights about therefolding mechanism of immunotoxin in the presence of anti-aggregators [4,5].