L. V. Dubey

Interaction of a tricationic meso-substituted porphyrin with guanine-containing polyribonucleotides of various structures

The interaction of a tricationic water-soluble meso-(N-methylpyridinium)-substituted porphyrin, TMPyP3+, derived from classic TMPyP4, with double-stranded poly(G)  ⋅  poly(C) and four-stranded poly(G) polyribonucleotides has been studied in aqueous buffered solutions, pH 6.9, of low and near-physiological ionic strengths in a wide range of molar phosphate-to-dye ratios (P/D). To clarify the binding modes of TMPyP3+ to biopolymers various spectroscopic techniques, including absorption and polarized fluorescence spectroscopy, Raman spectroscopy, and resonance light scattering, were used. As a result, two competitive binding modes were revealed. In solution of low ionic strength outside binding of the porphyrin to the polynucleotide backbone with self-stacking prevailed at low P/D ratios (P/D  <  3.5). It manifested itself by the substantial quenching of porphyrin fluorescence. Also the formation of large-scale porphyrin aggregates was observed near the stoichiometric binding ratio. The spectral changes observed at P/D  >  30 including emission enhancement were supposed to be caused by the embedding of partially stacked porphyrin J-dimers into the polymer groove. TMPyP3+ binding to poly(G) induced a fluorescence increase 2.5 times as large as that observed for poly(G)  ⋅  poly(C). In solution of near-physiological ionic strength the efficiency of external porphyrin binding was reduced substantially due to the competitive binding of Na+ ions with the polymer backbone. The spectroscopic characteristics of porphyrin bound to polynucleotides at different conditions were compared with those for free porphyrin.

Spectroscopic Studies on Binding of Cationic Pheophorbide-a Derivative to Model Polynucleotides

The binding of new cationic Pheophorbide-a derivative to double-stranded poly(A)-poly(U), poly(G)-poly(C) and four-stranded poly(G) was investigated in buffered solutions, pH6.9, of low ionic strength (2 mM Na+) by methods of absorption and polarized fluorescence spectroscopy in a wide range of molar phosphate-to-dye ratios (P/D). Two types of CatPheo-a binding to polynucleotides were revealed: (i) dye intercalation between the nucleic bases; (ii) cooperative binding with stacking of chromophores caused by electrostatic attraction of the dye to polynucleotide backbones. The last type is predominant for double-stranded polynucleotides. However in the case of four-stranded poly(G) containing sytem the external binding prevails only at low P/D, whereas under high P/D values the intercalative binding mechanism is mainly realized. Fluorescent techniques was revealed to be efficient for recognition of binding mode because of the dye emission increases upon chromophore intercalation and quenches strongly upon the outside binding accompanied with the dye stacking-association The spectroscopic characteristics of complexes formed were obtained.

Spectroscopic Studies on Binding of Porphyrin-Phenazine Conjugate to Four-Stranded Poly(G)

Binding of a novel cationic porphyrin–imidazophenazine conjugate, TMPyP3+–ImPzn, to four-stranded poly(G) was investigated in aqueous solutions of neutral pH under near physiological ionic conditions using absorption, polarized fluorescent spectroscopy and fluorescence titration techniques. In absence of the polymer the conjugate folds into stable internal heterodimer with stacking between the porphyrin and phenazine chromophores. Binding of TMPyP3+–ImPzn to poly(G) is realized by two competing ways. At low polymer-to-dye ratio (P/D < 6) outside electrostatic binding of the cationic porphyrin moieties of the conjugate to anionic polynucleotide backbone with their self-stacking is predominant. It is accompanied by heterodimer dissociation and distancing of phenazine moieties from the polymer. This binding mode is characterized by strong quenching of the conjugate fluorescence. Increase of P/D results in the disintegration of the porphyrin stacks and redistribution of the bound conjugate molecules along the polymer chain. At P/D > 10 another binding mode becomes dominant, embedding of TMPyP3+–ImPzn heterodimers into poly(G) groove as a whole is occurred.

Binding of Metallated Porphyrin-Imidazophenazine Conjugate to Tetramolecular Quadruplex Formed by Poly(G): a Spectroscopic Investigation.

The binding of telomerase inhibitor ZnTMPyP3+–ImPzn, Zn(II) derivative of tricationic porphyrin–imidazophenazine conjugate, to tetramolecular quadruplex structure formed by poly(G) was studied in aqueous solutions at neutral pH and near physiological ionic strength using absorption and polarized fluorescent spectroscopy techniques. Three binding modes were determined from the dependences of the fluorescence intensity and polarization degree for the porphyrin and phenazine moieties of the conjugate on molar polymer-to-dye ratio (P/D). The first one is outside electrostatic binding of positively charged porphyrin fragments to anionic phosphate groups of the polymer which prevails only at very low P/D values and manifests itself by substantial fluorescence quenching. It is suggested that the formation of externally bound porphyrin dimers occurs. The other two binding modes observed at high P/D are embedding of the ZnTMPyP3+ moiety into the groove of poly(G) quadruplex accompanied by more than 3-fold enhancement of the conjugate emission, and simultaneous intercalation of the phenazine fragment between the guanine bases accompanied by the increase of its fluorescence polarization degree up to 0.25. Thus Zn(II) conjugate seems to be promising ligand for the stabilization of G-quadruplex structures since porphyrin binding to poly(G) is strengthened by additional intercalation of phenazine moiety.