In novo synthesis of fluorescent arsenical dye and conjugation with Thioredoxin: Spectrometry Characterization

FEMIA, L.; TEMPRANA, C.F.; SANTOS, J.; GRASSELLI, M,; ALONSO, S.DEL V.

Río de Janeiro

Congreso; III Latin American Postgraduate Program f Biophysics Course. I Colloquium Brazil/Africa of Biophysics; 2010

It is well known arsenic compounds application on different cancer therapies, like promyelocitic acute leukemia. Hereby is proposed reduced arsanilic acid as an alternative arsenic drug, since its toxicity is lower than inorganic arsenic compounds. My thesis deals with protein/arsenic compound interaction. Protein chosen was Thioredoxin, since it has vicinal cysteins which are capable to form disulfure bridges with arsenic(III). In first place we have synthetized a novel fluorescent arsenic dye between arsanilic acid and Fluorescein Isothiocyanate (FITC). The reaction product has been characterized. Infrared analysis and mass spectrometry has been made, a majority peak was observed with a q/m ratio of 594. UV-VIS spectra and fluorescence shows the presence of a compound with mixed properties and atomic absorption measurements confirm arsenic presence in the fluorescent compound. In addition, exhibits a direct relationship between the amount of As, absorbance at 494 nm and fluorescence at 515 nm. Moreover, the interaction with Trx is evident in the absorption and fluorescence spectra comparable to the compound, as well as presence of As. The secondary and tertiary structure of Trx-compound is maintained when making a circular dichroism analysis of it. In addition, when the labeled protein is treated with dithiothreitol shows the elimination of fluorescence, visible absorption and presence of As. This, coupled with the fact that Trx incubated with FITC, alone, under the same reaction conditions does not incorpore the probe, evidence the attach it is through As group.