The Role of Ground State Complexation in the Electron Transfer Quenching of Methylene Blue Fluorescence by Purine Nucleotides

Abstract

The effect of three purine nucleotides on the fluorescence of methylene blue in aqueous buffer has been investigated. Guanosine-5'-monophosphate (GMP) and xanthosine-5'-monophosphate cause fluorescence quenching while adenosine-5'-monophosphate causes a red shift in the fluorescence maximum. All three nucleotides form ground state complexes with the nucleotides as indicated by absorption spectroscopy. The fluorescence changes at nucleotide concentrations less than 30 mM are best described by a static mechanism involving the formation of non-fluorescent binary and ternary complexes in competition with dimerization of the dye. Quenching of the fluorescence decay (tau = 368 ps) at high GMP concentrations (10-100 mM) occurs at the rate of diffusion. The mechanism of fluorescence quenching may involve electron transfer within the singlet excited dye-nucleotide complex although published values of the oxidation potentials of various purine derivatives would suggest that all three nucleotides should cause quenching. Evidence for electron transfer was obtained from flash photolysis experiments in which 100 mM GMP was found to cause the appearance of a long lived transient species absorbing in the region expected for semimethylene blue.

Photochemical Cleavage of DNA With Phosphorus(V)porphyrin Derivatives

“…The free energy change for oxidation of nucleic acid bases by the singlet excited EG2P(V)TPP was sufficient for the electron transfer, that is -0.99V for guanine and -0.59V for adenine. 23 The electron transfer of the triplet excited EG2P(V)TPP was directly confirmed by the transient absorption measurements under the same conditions as the fluorescence quenching measurements, where ground state interaction took place. At 15011s after laser pulse excitation, the triplet absorptions of the EG2P(V)TPP around 480nm (FlG.3, C) were observed in the presence of either poly(dG)poly(dC) or poly(dA)poly(dT) (FIG.3, (a), (b) ), but the decay processes are quite different in each case.…”

Probing the Interaction Between Fluorophores and DNA Nucleotides by Fluorescence Correlation Spectroscopy and Fluorescence Quenching†

“…The interactions of fluorescent dyes with nucleotides have been extensively studied for a large variety of coumarins, rhodamines, xanthene dyes and cyanines by means of fluorescence quenching experiments (18)(19)(20)(21)(22). The nucleoside guanosine has the lowest oxidation potential among the four DNA bases, and can quench many fluorophores via photoinducedelectron transfer (19,21,23,24). Quenching by other nucleotides has been reported in other cases as well (21).…”

Mechanistic and Kinetic Aspects of Photosensitization in the Presence of Oxygen †§

“…But in fact, dGuo quenches the singlet excited state of MB ϩ , so that this value must be corrected in order to take this process into account: ␥ ⌬ G ϭ 0.06 in the presence of 10 Ϫ3 M dGuo. Such an efficient quenching of the singlet state of MB ϩ by guanosine derivatives has already been reported (40) and an electron-transfer mechanism was clearly argued for this interaction, the back electron transfer of the formed radicals being practically complete (41). The incident light intensity was calibrated using the photooxygenation of furfurylic alcohol sensitized by 2 ϫ 10 Ϫ5 M MB ϩ in water, pH 6 (upper curve of Fig.…”