Estudo do efeito do veneno da cobra Crotalus basiliscus na reação de transferência de elétrons da mioglobina

Abstract

The effect of the venom of the Crotalus basiliscus snake on the electron transfer (ET) reaction of the myoglobin (Mb) protein was studied using a gold electrode modified with cysteine (Au/Cys) followed by the immobilization of the protein (Au/Cys/Mb). For that, the following techniques were used: cyclic voltammetry, surface plasmon resonance (SPR), absorption spectroscopy in the ultraviolet and visible regions (UV-Vis), circular dicroism (CD), and gel electrophoresis. The electrode reaction assigned to the FeII/III redox pair of Mb was observed at 0.09V vs Ag/AgCl being consistent with the native form of the protein. Upon immersion of the Au/Cys/Mb electrode in solution containing the venom of the Crotalus basiliscus snake, it was observed a decrease in the faradaic current. The deleterious effect of the venom on the electrode reaction of Mb was corroborated by the observation of the loss of the catalytic activity of the Au/Cys/Mb electrode towards the oxidation reaction of the ascorbic acid (AA). The successive mass increments observed by SPR upon the injections of cysteine (0.172 µg cm-2), myoglobin (0.410 µg cm-2) and venom (0.151 µg cm-2) indicated that the adsorbed molecules of cysteine and myoglobin were not desorbed from the gold surface upon exposure to the solution of the venom of the Crotalus basiliscus snake. Rather, the mass increase indicated the molecules of the venom were immobilized on the the Au/Cys/Mb electrode. Such proposition was reinforced by the electrophoresis results which showed a decrease in the mobility of Mb after mixing to the solution containing the snake venom. The electronic UV-Vis spectra did not show meaningful changes in the Soret band of Mb; a band sensitive to the level of protein denaturation. Likewise, the CD spectra obtained in the far-UV, near-UV and Soret regions did not present spectral profile changes indicating that the physical integrity of the protein was maintained after the exposure to the snake venom, i.e. the protein was not denaturated.