Photochemical formation of a fluorescent thymidine - pterin adduct in DNA

SANDRA ESTÉBANEZ RUIZ; CAROLINA LORENTE; M. LUISA MARÍN; MIGUEL A. MIRANDA; VIRGINIE LHIAUBET- VALLET; ANDRÉS H. THOMAS

Congreso; XIII Encuentro Latinoamericano de Fotoquímica y Fotobiología; 2017

UNSE, UNC, UNLP

Most solar radiation incident on the surface of Earth in the UV-range corresponds to wavelenghtsbetween 320 -400 nm (UV-A). This fraction of radiation is poorly absorbed by the DNAbiomacromolecule, and therefore it is responsible for only low amounts of directly formed photolesions. Nevertheless, UV-A radiation acts indirectly by photosensitized reactions and is recognized as a class I carcinogen [1]. A photosensitized reaction is a photochemical modification occurring in a molecular entity as a result of the initial absorption of radiation by a photosensitizer [2]. Pterins belong to a family of heterocyclic compounds present in a wide range of living systems and participate in relevant biological functions. Under UV-A excitation, pterins can fluoresce, undergo photooxidation and generate reactive oxygen species (ROS) [3]. In the presence of oxygen, pterin (Ptr) act as photosensitizer through type I (electron abstraction) and/or type II (1O2-mediated oxidation) mechanisms [4-5].The photosensitized degradation of the pyrimidine nucleotide thymidine 5?-monophosphate(dTMP) by Ptr under anaerobic conditions generates an adduct, where the pterinic moiety is attached to the nucleobase. The spectroscopic properties of the adduct are similar to those of Ptritself [6].The main objective of this work was to test the photodegradation of an oligonucleotide chain in the presence of Ptr and UV-A. Single stranded oligonucleotide of dTMP with the sequence 5 ́-d(TTTTT)-3 ́ (dT5) was exposed to UV-A radiation in the presence of Ptr in neutral aqueous solutions at room temperature, under different experimental conditions. The samples were analyzed by UV-Vis spectrophotometry, HPLC coupled to a photodiode array and fluorescence detector and UPLC coupled to a mass spectrometry system.Chromatographic peaks corresponding to both dT5 and Ptr decreased with irradiation time. Several products presented absorbance in the UV-A region, and their absorption spectra showed a band centered approximately at 340 nm, which is similar to the typical low-energy band of Ptr. Moreover, some of these products were fluorescent and emitted at 450 nm when excited at 350 nm, which is compatible with the emission properties of Ptr. Products were analyzed by mass spectrometry, and the mass spectra showed a group of signals with m/z 808.65, which correspond to a di-charged ion of a compound bearing both of the photosensitizer and the oligonucleotide moieties. Thus, a covalent bond between Ptr and dT5 occurs in anaerobic conditions.