Influence of Ionizing Radiation on Spontaneously Formed Aggregates in Proteins or Enzymes Solutions.

We have shown that many proteins and enzymes (ovalbumin, β-lactoglobulin, lysozyme, insulin, histone, papain) undergo concentration-dependent reversible aggregation as a result of the interaction of the studied biomolecules. Moreover, irradiation of those protein or enzyme solutions under oxidative stress conditions results in the formation of stable soluble protein aggregates. We assume that protein dimers are mainly formed. A pulse radiolysis study has been made to investigate the early stages of protein oxidation by N 3 • or •OH radicals. Reactions of the N 3 • radical with the studied proteins lead to the generation of aggregates stabilized by covalent bonds between tyrosine residues. The high reactivity of the •OH with amino acids contained within proteins is responsible for the formation of various covalent bonds (including C–C or C–O–C) between adjacent protein molecules. In the analysis of the formation of protein aggregates, intramolecular electron transfer from the tyrosine moiety to Trp• radical should be taken into account. Steady-state spectroscopic measurements with a detection of emission and absorbance, together with measurements of the dynamic scattering of laser light, made it possible to characterize the obtained aggregates. The identification of protein nanostructures generated by ionizing radiation using spectroscopic methods is difficult due to the spontaneous formation of protein aggregates before irradiation. The commonly used fluorescence detection of dityrosyl cross-linking (DT) as a marker of protein modification under the influence of ionizing radiation requires modification in the case of the tested objects. A precise photochemical lifetime measurement of the excited states of radiation-generated aggregates is useful in characterizing their structure. Resonance light scattering (RLS) has proven to be an extremely sensitive and useful technique to detect protein aggregates. [ABSTRACT FROM AUTHOR]