Influence of reactive radicals OH and Esup(-)sub(AQ) in irradiation of protein in aqueous solutions

Abstract

Gamma irradiation of aqueous proteins solutions promotes structural modifications. This is mainly caused by OH • and eaq activity, reactive species produced by water radiolys. The objective of this study was to investigate the role of OH • and eaq in conformation changes observed in proteins irradiated in aqueous solutions. Substances that react specifically with OH • or eaq producing stable products were used in order to study each reactive species’ function: nitrate for eaq and t-butilic alcohol for OH • . Crotamine was used as test peptide. It is composed of 42 aminoacids, which are strongly reticulated by three disulfide bonds. This confers it a very compact conformation leading to high temperatures resistance. Crotamine also has a biological effect: mice hind limb distention. Samples of crotamine in aqueous solution were irradiated in order to analyze the activity of OH • and eaq. Some contemned t-butilic alcohol and others, sodium nitrate; the results were compared to control samples with no neutralizing substances. To verify possible protein conformation change due to sodium nitrate or t-butilic alcohol addition, samples with different concentrations of these substances were prepared, kept without irradiation and their results were compared to only protein solution. Samples were also submitted to a biological test Samples were submitted to SDS-PAGE, which did not show any alteration in those samples with t-butilic alcohol or sodium nitrate. In addition, the samples had their absorbance spectrum of 200 nm to 360 nm scanned which showed less conformation change in irradiated with t-butilic then only protein one. Irradiated samples had less biological effect than non-irradiated ones.