Blue autofluorescence in protein aggregates “lighted on” by UV induced oxidation

• Published in: Biochimica et biophysica acta. Proteins and proteomics 2019-11, Vol.1867 (11), p.140258, Article 140258
• Main Authors: Fricano, A., Librizzi, F., Rao, E., Alfano, C., Vetri, V.
• Format: Article
• Language: English
• Subjects: Amyloid autofluorescence; Amyloid beta-Peptides - chemistry; Dityrosine formation; Fluorescence; Fluorescence lifetime imaging; Humans; Oxidation-Reduction - radiation effects; Oxidative stress; Protein Aggregates - radiation effects; Ubiquitin; Ultraviolet Rays
• ISSN: 1570-9639; 1878-1454
• DOI: 10.1016/j.bbapap.2019.07.011

Oxidation of amino acid side chains in protein structure can be induced by UV irradiation leading to critical changes in molecular structure possibly modifying protein stability and bioactivity. Here we show, by using a combination of multiple spectroscopic techniques and Fluorescence Lifetime Imaging, that UV-light exposure induces irreversible oxidation processes in Ubiquitin structure. In particular, the growth of a new autofluorescence peak in the blue region is detected, that we attribute to tyrosine oxidation products. Blue autofluorescence intensity is found to progressively increase also during aggregation processes leading to the formation of aggregates of non-amyloid nature. Significantly, analogous spectral modifications are found in amyloid fibrils from human insulin and Amyloid-β peptide grown under UV exposure. Experimental results reveal a substantial overlap between the fluorescence signal here attributed to tyrosine oxidation and the one referred in literature as “Amyloid autofluorescence”. These findings clearly represent a caveat about the specificity of the blue fluorescence peak measured for amyloids, especially when grown in conditions in which tyrosine residues may be oxidized. Moreover, our results once again highlight the close link between the formation of amyloid aggregates and protein damage resulting from oxidative stress, as these neurotoxic aggregate species are found to contain damaged residues. [Display omitted] •UV light induces Ubiquitin oxidation.•UV light modifies observed protein thermal aggregation kinetics.•Protein damages can be induced by standard instrumentations.•Amyloid intrinsic autofluorescence overlaps with tyrosine oxidation products signal.