Ubiquitin?Pterin fluorescent adduct formation under UV-A radiation

REID, LARA O.; DANTOLA, MARIA LAURA; MARIN, MARIA LUISA; LHIAUBET-VALLET,VIRGINIE; THOMAS, ANDRÉS H.

La Plata

Congreso; XLVII Reunión Anual de la Sociedad Argentina de Biofísica; 2018

Sociedad Argentina de Biofísica

Different compounds are able to inducephotosensitivity as a result from exposure to certain molecules and light;these includes phototoxic and photoallergic reactions. The photoallergy normallyinvolves a covalent binding between proteins and photosensitizer agents leadingto the formation of a complete photoantigen, which may trigger ahypersensitivity reaction due to a cell-mediated immune response.Pterins belong to a family of heterocycliccompounds present in a wide range of living systems and this compounds are able to photosensitize damage in proteins, DNAand their components by Type I (electron ?transfer) and Type II (singletoxygen) mechanisms.1,2Therefore, given thebiological and medical relevance of the photosensitizing properties of pterins,the aim of this work isto study if pterin (Ptr), the parent and unsubstitutedcompound of oxidized pterins, is able to generate photoadducts with proteins and establish its photoallergicpotencial. For this study, aqueous acidic solution of Ubiquitin (Ub) and Ptrwere irradiated (lex=350 nm) at room temperature. Ub wasused as a model protein given that is a small (8.5 kDa) regulatory protein, whichhas only one Tyrosine (Tyr) residue and none Tryptophan residue.The irradiated solution were analyzed by UV/visible spectrophotometry,HPLC, fluorescence spectroscopy and SDS-PAGE.Under UV-A radiation Ptr is able to form an adduct with Ub, and thisreaction is much more efficient in the absence of O2. Thespectroscopic analysis reveals that the emission and the excitation spectrumare similar to those corresponding to Ptr, as well as the fluorescencelifetime. On the other hand, as a consequence of the photosensitized process,the protein suffers oligomerization mediated by Tyr dimers, and also afragmentation can occur, which is dependent of the oxygen concentration in theatmosphere. 1-Lorente C., ThomasA. H., Acc. Chem. Res., 39, 395, 2006.2-Reid L. O., et al, Biochemistry, 55(34), 4777, 2016.