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The binding mode of cationic porphyrin ( trans -BMPyP) with poly[d(G-C) 2 ] and poly[d(A-T) 2 ] was examined according to the site of the periphery cationic methyl pyridine ion of the cationic porphyrin ( o-, m-, p- ) as well as the possibility of a B-Z transition depending on the binding modes by measuring the absorption spectrum and circular dichroism (CD). The negative band found in the soret region showed the intercalation mode of m- and p-trans- BMPyP-poly[d(G-C) 2 ] to the DNA base pairs, but no B-Z transition was induced. On the other hand, the distinctive bisignate band found in the soret region of the CD spectrum for m- and p-trans- BMPyP-poly[d(A-T) 2 ] suggests that m- and p-trans- BMPyP have an effective extensive stacking-based binding mode along with the skeleton of poly[d(A-T) 2 ], wherein the B-Z transition was induced through extensive stacking. The difference in binding mode was attributed to the difference in the molecular structure depending on the site of the periphery cationic methyl pyridine ion in the cationic porphyrin. In other words, o-trans- BMPyP is nonplanar because of the steric hindrance of the cationic methyl pyridine ion at the o- site. In contrast, m- and p-trans- BMPyP are planar, but not all porphyrins with a planar structure undergo the B-Z transition. In conclusion, a B-Z transition is induced if the structure of a porphyrin is planar and the binding mode allows the porphyrins to be stacked effectively along the DNA skeleton, not in a binding mode where the porphyrin is intercalated to the DNA.Communicated by Ramaswamy H. Sarma.

The spectral properties of meso -tetrakis ( N -methylpyridinium-4-yl)porphyrin (TMPyP) in the presence of parallel and antiparallel G-quadruplexes formed from a thrombin-binding aptamer G-quadruplex (5'-G 3 T 2 G 3 TGTG 3 T 2 G 3 ) were investigated in this study. Red shift and hypochromism in the Soret absorption band of TMPyP were observed after binding to both parallel and antiparallel G-quadruplexes. The extent of changes in the absorption spectra were similar for both conformers. No circular dichroism spectrum was induced in the Soret region for both parallel and antiparallel G-quadruplexes. This is suggest that there is no or very weak interaction between electric transitions of nucleobases and porphyrin molecule. The accessibility of the neutral quencher I 2 to the G-quadruplex-bound TMPyP was similar for both parallel and antiparallel G-quadruplexes. All these observations suggest that TMPyP was bound at the outside of the quadruplexes, and conceivably interacted with the phosphate group via a weak electrostatic interaction.Communicated by Ramaswamy H. Sarma.

The binding modes of a pyrene-porphyrin dyad, (1-pyrenyl)-tris(N-methyl-p-pyridino)porphyrin (PyTMpyP), to various DNAs (calf thymus DNA (Ct-DNA), poly[d(G-C)2], and poly[d(A-T)2]) have been investigated using circular dichroism and linear dichroism measurements. Based on the polarization spectroscopic results, it can be shown that the pyrenyl and porphryin planes are skewed to a large extent for PyTMPyP in an aqueous environment and in the binding site of poly[d(G-C)2]. In this complex, a photoinduced electron transfer (PET) process between the pyrenyl and porphyrin moieties occurs. On the other hand, PET was not observed in the PyTMPyP-poly[d(A-T)2] complex, whereas the fluorescence intensity of TMPyP was enhanced. The molecular planes of the pyrene and porphyrin moieties are almost parallel in the poly[d(A-T)2] and Ct-DNA adducts. Moreover, the generation of 1O2 species occurs only for the PyTMPyP-Ct-DNA and PyTMPyP-poly[d(A-T)2] complexes. We discuss the photophysical properties of PyTMPyP which are attributed to the binding patterns and the sequence of DNA bases.

The binding modes of a pyrene-porphyrin dyad, (1-pyrenyl)-tris(N-methyl-p-pyridino)porphyrin (PyTMpyP), to DNA and its photophysical properties have been investigated using various spectroscopic techniques. The circular dichroism (CD) spectrum of PyTMpyP bound to DNA (PyTMpyP-DNA) showed one negative and two positive bands in the Soret region. The CD signal in the pyrene absorption region was positive. The shape of the CD spectrum does not support an intercalative binding mode of TMpyP, which would typically afford a negative CD band in the absence of the pyrene moiety. Linear dichroism (LD) experiments revealed a very small signal in the Soret region, which also challenges the intercalation of TMpyP into DNA. Upon excitation of the pyrene moiety, the emission intensity of porphyrin in aqueous solution was quenched due to a photoinduced electron transfer (PET) process between the pyrenyl and porphyrin moieties. On the other hand, the emission of porphyrin was markedly enhanced upon binding to DNA, as the PET process from the excited pyrene moiety to TMpyP was suppressed when bound to DNA. The PET process occurs in the timescale of 65 ps, and could be detected by femtosecond transient absorption spectroscopic methods. Two fluorescence decay times were observed for PyTMpyP in aqueous solution (0.78 and 4.8 ns). Both decay times increased upon binding to DNA owing to environment and/or conformational changes in PyTMpyP. The driving force (ΔG) of the PET process was evaluated under conditions of minor and major groove binding. The PET process and photophysical properties of the PyTMpyP dyad were concluded to be influenced by the binding mode.

In this study, the binding mode of porphyrin-free single-stranded poly[d(AT)] and trans-BMPyP was observed in the Z-form trans-BMPyP-poly[d(A-T) 2 ] complex induced by extensive stacking depending on the temperature and concentration through circular dichroism (CD). The Z-form trans-BMPyP-poly[d(A-T) 2 ] complex (R = 0.30) retained the Z-form DNA structure at a low temperature (20 °C) by the trans-BMPyP molecules. When the temperature was increased to 60 °C, the DNA was almost unfolded as a single-stranded poly[d(AT)], but the extensive stacking binding mode of trans-BMPyP was maintained and the shape of the porphyrin Soret band was symmetrically changed in comparison with the shape of the Z-form DNA. However, when the temperature was raised to 80 °C, the extensive stacking binding mode of trans-BMPyP was also unfolded almost completely. The binding mode of the trans-BMPyP-single-stranded poly[d(AT)] complex was very similar to the already known binding mode of porphyrins and a double-stranded DNA. The binding mode was dependent on the concentration ratio ([porphyrin]/[DNA]): a monomeric binding mode at a concentration ratio of 0.04, a moderate groove binding mode at a concentration ratio between 0.08 and 0.16, and extensive stacking at a concentration ratio between 0.20 and 0.30. The same result was obtained when the temperature of the Z-form DNA (R = 0.30) was increased to 60 °C. However, those binding modes were not found in cis-BMPyP, which was because, in the extensive stacking of trans-BMPyP along the DNA skeleton, the distance between the two positive methylpyridine ions at the trans site and thymine, one of the DNA bases, is decreased, creating a much more hydrophobic environment. In addition, the poly AT sequences found from the CD spectra for the binding of trans-BMPyP-poly[d(A-T) 2 ] and trans-BMPyP-poly[d(AT)] (R = 0.30) showed that both of them underwent effective extensive stacking and that the chirality of extensive stacking was dependent on the form of DNA.

trans-BMPyP induced the B-Z transition for alternated AT oligonucleotides as it was evident by inversed CD spectrum. The transition occurred simultaneously with appearance of the extensive stacking of porphyrin. Complete B-Z transition required at least 14 base-pairs long. Insertion of one or two GC base pairs prevented the B-Z transition., (Copyright © 2016. Published by Elsevier B.V.)

The thermal stability of the G-quadruplex formed from the thrombin-binding aptamer, 5'G2T2G2TGTG2T2G2, in which the guanine (G) base at the central loop was replaced with an adenine (A) or inosine (I) base, was examined to determine the role of the central G base in stabilizing the quadruplex. Replacement of the central G base by the I base resulted in a slight decrease in thermal stability. On the other hand, the stability of the G-quadruplex decreased to a significant extent when it was replaced with the A base. The optimized structure of the G-quadruplex, which was obtained by a molecular dynamic simulation, showed that the carbonyl group of the C5 position of the central G base could form hydrogen bonds with the G1 amine group at the C7 position on the upper G-quartet. This formation of a hydrogen bond contributes to the stability of the G-quadruplex. The spectral property of meso-tetrakis(N-methylpyridium-4yl)porphyrin (TMPyP) associated with the G-quadruplex was characterized by a moderate red shift and hypochromism in the absorption spectrum, a positive CD signal, and two emission maxima in the fluorescence emission spectrum, suggesting that TMPyP binds at the exterior of the G-quadruplex. Spectral properties were slightly altered when the G base at the central loop was replaced with A or I, while the fluorescence decay times of TMPyP associated with the G-quadruplex were identical. Observed spectral properties removes the possibility of intercalation binding mode for TMPyP. TMPyP binds at the exterior of the quadruplex. Whether it stacks on the central loop or binds at the side of the quadruplex is unclear at this stage., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Typical CD spectrum of the right-handed poly[d(A-T)2] was reversed when trans-bis(N-methylpyrimidium-4-yl)diphenyl porphyrin (trans-BMPyP) was bound, suggesting that the helicity of the polynucleotide was reversed to the left-handed form. The formation of the left-handed Z-form poly[d(A-T)2] was confirmed by (31)P NMR, in which a single (31)P peak of B-form poly[d(A-T)2] was split into two peaks, which is similar to the conventional B-Z transition of poly[d(G-C)2] induced by the high ionic strength. The observed B-Z transition is unique for poly[d(A-T)2]. The other polynucleotides, including poly[d(G-C)2], poly(dG)·poly(dC) and poly(dA)·poly(dT) remained as the right-handed form in the presence of the same porphyrin. This observation suggests that the porphyrin array that was formed along the poly[d(A-T)2] provides a template to which left-handed poly[d(A-T)2] is associated with an electrostatic interaction.

The effects of the central metal ion on complex formation between meso-tetrakis(N-methylpyridium-4-yl)porphyrin (TMPyP) and the thrombin-binding aptamer G-quadruplex, 5'G2T2G2TGTG2T2G2, were examined in this study. The central metal ions were vanadium and zinc. At a [porphyrin]/[G-quadruplex] ratio of less than one, the absorption and CD spectra were unaffected by the mixing ratio for all three porphyrins, suggesting that the binding mode is homogeneous. Relatively small changes in the absorption spectrum when forming the complexes with the G-quadruplex, the positive CD signal, and the large accessibility of the I(-) quencher, suggested that all these porphyrins are not intercalated between the G-quartet. Stabilization of the G-quadruplex by ZnTMPyP was most effective. The effect of VOTMPyP on G-quadruplex stabilization was moderate, whereas TMPyP slightly destabilized G-quadruplex. From this observation, the involvement of the ligation of one G-quartet component to the central metal ion in G-quadruplex stabilization by metallo-TMPyP is suggested., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The photoreduction of water-soluble cationic manganese(III) meso-tetrakis(1-methylpyridium-4-yl)porphyrin (Mn(III)(TMPyP)(4+)) bound to a synthetic polynucleotide, either poly[d(A-T)2] or poly[d(G-C)2], was examined by conventional absorption and circular dichroism (CD) spectroscopy, transient absorption, and transient Raman spectroscopy. Upon binding, Mn(III)(TMPyP)(4+) produced a positive CD signal for both polynucleotides, suggesting external binding. In the poly[d(A-T)2]-Mn(III)(TMPyP)(4+) adduct case, an interaction between the bound porphyrin was suggested. The transient absorption spectral features of Mn(III)(TMPyP)(4+) in the presence of poly[d(A-T)2] and poly[d(G-C)2] were similar to those of the photoreduced products, Mn(II)(TMPyP)(4+), whereas Mn(III)(TMPyP)(4+) in the absence of polynucleotides retained its oxidation state. This indicated that both poly[d(A-T)2] and poly[d(G-C)2] act as electron donors, resulting in photo-oxidized G and A bases. The transient Raman bands (ν2 and ν4) that were assigned to porphyrin macrocycles exhibited a large downshift of ~25 cm(-1), indicating the photoreduction of Mn(III) to Mn(II) porphyrins when bound to both polynucleotides. The transient Raman bands for pyridine were enhanced significantly, suggesting that the rotation of peripheral groups for binding with polynucleotides is the major change in the geometry expected in the photoreduction process. These photoinduced changes do not appear to be affected by the binding mode of porphyrin.

The binding modes of cationic porphyrins, namely M-meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (MTMPyP, where M = free base, Cu(II), Ni(II), Co(III), Mn(III), V(IV)═O, and Ti(IV)═O), to native DNA were systematically investigated by polarized light spectroscopies, viz. circular and linear dichroism spectroscopy (CD and LD, respectively). At low [porphyrin]/[DNA] ratio, planar porphyrins including TMPyP, CuTMPyP, and NiTMPyP exhibited a negative CD signal in the Soret region and large negative wavelength-dependent LD(r) signal which is characteristics of their intercalative binding mode. In the intercalation pocket, the molecular plane of porphyrin is skewed to a large extent as it was judged from a large wavelength-dependency of the LD(r) magnitude in the Soret region. As the mixing ratio was increased, a bisignate CD spectrum became apparent for all of the planar porphyrins, while the shape of LD(r) remained the same, indicating the coupling of the electric transition moments of the intercalated porphyrins. Thus, coupling can occur between the porphyrins when they are separated at least two DNA base-pairs according to the nearest neighboring site exclusion model. This coupling interaction was the weakest for NiTMPyP. The porphyrins with axial ligands, namely VOTMPyP, TiOTMPyP, MnTMPyP, and CoTMPyP, bind to the exterior of the DNA at a low [porphyrin]/[DNA] ratio, which is indicated by the positive CD signal in the Soret region. The absorption and LD spectra in the Soret region were treated as the sum of two transitions to calculate the angle between each of the electric transition moments of the porphyrin, i.e., the B(x) and B(y) transitions relative to the local DNA helix axis. The angle of one of the two electric transitions of the porphyrin is in the range of 56°∼59°, whereas the other is in the range of 59°∼65°. Increasing the porphyrin density resulted in the appearance of an interaction between the bound porphyrins, which was indicated by the distorted CD spectra. In contrast with the planar porphyrins, which are parallel to each other in the intercalation pocket when they interact by themselves, the molecular plane of the porphyrin with the axial ligand tilts more toward the local DNA helix axis as the population of porphyrins was increased. CoTMPyP exhibited complicated CD and LD spectra at high [porphyrin]/[DNA] ratios that were impossible to explain by the presence of two porphyrin species: those with a monomeric external binding and electric coupling. Since the CoTMPyP-DNA complex started to aggregate at relatively low [porphyrin]/[DNA] ratios, the complication may due to the involvement of extensive porphyrin aggregation.

Zn(II) meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (ZnTMPyP) produced a unique linear dichroism (LD) spectrum when forming a complex with Z-form poly[d(G-C)(2)]. The spectrum was characterized by a large positive wavelength-dependent LD signal in the Soret absorption region. The magnitudes of LD in both the DNA and Soret band increased as the [porphyrin]/[DNA base] ratio increased and were larger by 20-40 times compared to the negative LD of the ZnTMPyP bound to B-form poly[d(G-C)(2)] and poly[d(A-T)(2)]. The angles calculated from LD were respectively 49° and 42° for B(x) and B(y) transitions of the porphyrin with respect to the local helix axis of Z-form poly[d(G-C)(2)]. The appearance of the unique LD spectrum for the Z-form poly[d(G-C)(2)] complex was accompanied by a bisignate circular dichroism spectrum in the Soret region, whose magnitude was proportional to the square of the porphyrin concentration, suggesting a stacking interaction between Z-form poly[d(G-C)(2)]-bound ZnTMPyP with other bound ZnTMPyP. From these observations, a conceivable binding mode of ZnTMPyP to Z-form poly[d(G-C)(2)] complex was proposed, in which ZnTMPyP binds at the major groove or across the groove. In contrast with Z-form poly[d(G-C)(2)], ZnTMPyP binds to poly[d(A-T)(2)] in a monomeric manner with the angles of 57° and 59° for the two porphyrin's transition moments with respect to the local polynucleotide helix axis. The polarized spectral properties of ZnTMPyP bound to B-form poly[d(G-C)(2)] coincide with the intercalated nonmetallic TMPyP, namely, a negative CD signal in the Soret band and a negative wavelength-dependent reduced LD signal, with a magnitude larger than that in the DNA absorption region in spite of its axial ligands.

Binding geometry to DNA and the efficiency as a donor for energy transfer of various metallo- and nonmetallo-porphyrins were investigated mainly by polarized light spectrscopy and fluorescence measurements. Planar porphyrins including nonmetallo meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (TMPyP), CuTMPyP, and NiTMPyP produced large red-shift and hypochromism in absorption spectrum and a negative circular dichroism (CD) in the Soret band suggesting that these porphyrins intercalate between DNA base-pairs as expected. In the intercalation pocket, the molecular plane of these porphyrins tilts to a large extent. From a linear dichroism (LD) study, the angle between the two electric transition moments in the Soret band were 16°, 12°, and 11° for TMPyP, NiTMPyP, and CuTMPyP, respectively. On the other hand, porphyrins with axial ligands namely, VOTMPyP, TiOTMPyP, and CoTMPyP, produced a positive CD signal in the Soret band. Hyperchromism and less red-shift were apparent in the absorption spectrum. These observations indicated that the porphyrins with an axial ligand bind outside of the DNA. The angles of both the B(x) and B(y) transition with respect to the local DNA helix were 39°~46° for all porphyrins. From these results, the conceivable binding site of porphyrins with axial ligands is suggested to be the minor groove. All porphyrins were able to quench the fluorescence of intercalated ethidium. Strong overlap between emission spectrum of ethidium and the absorption spectrum of porphyrins when they simultaneously bound to DNA was found suggesting the mechanism behind energy transfer is, at least in part, the Förster type resonance energy transfer (FRET). The minimum distance in base pairs between ethidium and porphyrin required to permit the excited ethidium to emit a photon was the longest for CoTMPyP being 17.6 base pairs and was the shortest for CuTMPyP and NiTMPyP at 8.0 base pairs. The variation in the distance was almost proportional to the extent of the spectral overlap, the common area under emission spectrum of ethidium and absorption spectrum of porphyrin, supporting the FRET mechanism, whereas the effect of the orientation factor which was considered by relative binding geometry was limited., (© 2012 American Chemical Society)

At high porphyrin densities, the effects of the number and position of the positive charges of the periphery ring on the stacking of the porphyrin on poly[d(A-T)(2)] was investigated using polarized spectroscopy, including circular and linear dichroism (CD and LD, respectively). The CD spectrum of meso-tetrakis(N-methylpyridinium-4-yl)porphyrin(TMPyP) consisted of two positive bands in the Soret absorption region at low [porphyrin]/[DNA base] ratios (R ratios) and changed to two distinguishable categories of the bisignate CD spectrum with increasing R ratio. These CD spectra were attributed to the monomeric groove binding, and the moderately and extensively stacked TMPyPs. In contrast, trans-bis(N-methylpyridinium-4-yl)porphyrin (trans-BMPyP) dominantly produced a CD spectrum that corresponded to the extensive stacking, except at the lowest R ratio that was used in this work (R = 0.04). However, for cis-bis(N-methylpyridinium-4-yl)porphyrin (cis-BMPyP), the intensity of the apparent bisignate CD signal was too small to assign it to the extensive stacking. Moreover, the shape of the CD spectrum in the DNA absorption region showed that the conformation of poly[d(A-T)(2)] was retained, in contrast to the extensively stacked TMPyP and trans-BMPyP. In the extensively stacked TMPyP- poly[d(A-T)(2)] assembly, the large negative LD signal in the Soret band was observed suggesting that the direction of the molecular planes of TMPyP was close to perpendicular with respect to the orientation axis (flow axis). In contrast, the LD spectrum of the trans-BMPyP-poly[d(A-T)(2)] complex produced positive LD signal in the same wavelength region, suggesting the orientation of the molecular plane was nearly parallel relative to the flow direction. Surprisingly, the LD signal in the DNA absorption region for both of the porphyrins was positive. Therefore, the helix axis of the DNA was near perpendicular relative to the flow direction in the porphyrin-polynucleotide assembly.

The circular and linear dichroism (CD and LD) spectral properties of the meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (TMPyP)-DNA complex at a [porphyrin]/[DNA] ratio below 0.015 showed that TMPyP intercalates between DNA base pairs. Contrarily, when cis-bis(N-methylpyridinium-4-yl)porphyrin (BMPyP) is associated with DNA, no CD spectrum was induced and a bisignate LD spectrum was observed. These spectral properties of both the TMPyP and BMPyP were essentially retained when the minor groove of the DNA was saturated with 4',6-diamidino-2-phenylindole (DAPI). The fluorescence of the DNA-bound DAPI was effectively quenched by BMPyP and TMPyP. The quenching by BMPyP can be described through a pure static mechanism while TMPyP quenching produced an upward bending curve in the Stern-Volmer plot. Quenching efficiency was by far greater than predicted by the "sphere of action model", suggesting that the DNA provides some additional processes for an effective energy transfer.

The effect of the number and position of the positive charges on porphyrin with respect to the mode of binding to poly[d(G-C)2] and poly[d(A-T)2] were investigated by absorption and polarized spectroscopy, including circular and linear dichroism (CD and LD). Meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (TMPyP), which possesses four positive charges on the periphery pyridinium rings, produces a negative CD and wavelength-independent reduced LD (LDr) spectra in the Soret absorption region when it associates with poly[d(G-C)2]. These spectral characteristics have been considered as diagnostic for intercalation. In contrast, both trans- and cis-bis(N-methylpyridinium-4-yl)diphenylporphyrin (trans- and cis-BMPyP), where the number of positive charges was reduced to two, multisignate CD and strong wavelength-dependence of the LDr spectra were observed, indicating that these porphyrins do not intercalate. Therefore, four positive charges are required for TMPyP intercalation. When associated with poly[d(A-T)2], trans-BMPyP exhibited a positive CD signal at a low [porphyrin]/[nucleobase] ratio with the appearance of a bisignate CD upon increase of the mixing ratio, suggestive of binding at the groove of the double helix at low mixing ratios, and stacking at increasing mixing ratios. Conversely, no monomeric binding was evident in the bis-BMPyP bisignate CD spectrum; hence, only the stacking mode was found for cis-BMPyP, even at the lowest [porphyrin]/[nucleobase] ratio, suggesting the importance of the position of the positive charges in determining monomeric groove binding or stacking. The binding geometries of trans- and cis-BMPyP were similar when associated with poly[d(A-T)2], as determined from the similar LDr spectrum. When associated with DNA, TMPyP exhibited similar spectral properties with that of the TMPyP-poly[d(G-C)2] complex, indicating intercalation of TMPyP between the DNA base pairs. Conversely, CD and LDr characteristics of both trans- and cis-BMPyP-DNA complexes resembled those that complexed with poly[d(A-T)2] at a high [porphyrin]/[DNA] ratio, suggesting that both porphyrins were stacked along the DNA stem.

The time-dependent binding mode of a porphyrin dimer to poly[d(G-C)2] and poly[d(A-T)2] was investigated by spectroscopic methods including absorption and circular and linear dichroism (CD and LD) spectroscopy. Immediately after mixing with poly[d(G-C)2], the porphyrin dimer exhibited red-shift and hypochromism in the absorption spectrum and negative CD and LD spectra. With further red-shift in absorption, the CD and LD magnitude in the Soret region became increasingly negative over time. After it was stabilized, the magnitude of the reduced LD (LDr) in the Soret region was larger than that in the DNA absorption region, indicating that the second porphyrin was also intercalated. Following the rapid intercalation of the first porphyrin, the very slow intercalation of the second followed with first-order kinetics. In the poly[d(A-T)2] case, a bisignate CD spectrum was observed in the Soret region suggesting stacking of the porphyrins. The small alteration in the CD spectrum and increased absorbance, which followed the initial rapid spectral change, was of the second order. This alteration in the spectral properties was attributed to the conformational change of poly[d(A-T)2] near the binding site because the overall shape of the CD spectrum was conserved in spite of the changes in the absorption spectrum.

The binding site and the geometry of Co(III)meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (CoTMPyP) complexed with double helical poly(dA).poly(dT) and poly(dG).poly(dC), and with triple helical poly(dA).[poly(dT)](2) and poly(dC).poly(dG).poly(dC)(+) were investigated by circular and linear dichroism (CD and LD). The appearance of monomeric positive CD at a low [porphyrin]/[DNA] ratio and bisignate CD at a high ratio of the CoTMPyP-poly(dA).poly(dT) complex is almost identical with its triplex counterpart. Similarity in the CD spectra was also observed for the CoTMPyP-poly(dG).poly(dC) and -poly(dC).poly(dG).poly(dC)(+) complex. This observation indicates that both monomeric binding and stacking of CoTMPyP to these polynucleotides occur at the minor groove. However, different binding geometry of CoTMPyP, when bind to AT- and GC-rich polynucleotide, was observed by LD spectrum. The difference in the binding geometry may be attributed to the difference in the interaction between polynucleotides and CoTMPyP: in the GC polynucleotide case, amine group protrude into the minor groove while it is not present in the AT polynucleotide.

The binding modes of the free-base meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (TMPyP) complexed with [d(AT)n]2 oligonucleotides (where n=3-8, corresponding to 6 to 16 AT base pairs) were studied by circular dichroism (CD). When associated with the shortest oligonucleotide, ([d(AT)3]2), a bisignate CD spectrum was produced in the Soret absorption region at the mixing ratio between 2.0 and 0.25, corresponding to one TMPyP per 0.5 to 4 oligonucleotides. Apparent bisignate CD was attributed to a stacked TMPyP along the DNA. On the other hand, when the oligonucleotide length reaches one helical turn or longer, ([d(AT)n]2, n=6,7,8), TMPyP exhibited a positive CD signal, that corresponds to the monomeric groove binding mode, at the mixing ratio below 1.0 (one TMPyP per oligonucleotide). As the mixing ratio increases, the CD signal was best accounted for by the sum of the stacked and groove-binding TMPyP. At the intermediate oligonucleotide length ([d(AT)n]2, n=4,5), the CD spectrum appeared to be the sum of the stacked and groove binding TMPyP at all mixing ratios. Therefore, it is conclusive that the full dispersion of TMPyP requires at least one helical turn of the AT sequence at a mixing ratio below 1.0. Further increase of the mixing ratio resulted in the stacking of TMPyP even at the long oligonucleotides.

The spectral properties of meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (TMPyP) bound to poly(dA).poly(dT) and poly[d(A-T)(2)] in the presence and in the absence of 4',6-diamidino-2-phenylindole (DAPI) have been studied. DAPI fits deeply into the minor groove of both poly(dA).poly(dT) and poly[d(A-T)(2)], and TMPyP is also situated at the minor groove. The nature of the absorption, circular dichroism (CD), and flow linear dichroism (LD) spectra of the TMPyP-poly(dA).poly(dT) and -poly[d(A-T)(2)] complexes in the Soret band is essentially unaffected whether the minor groove is blocked by DAPI or not, although small variations been noticed in the presence of DAPI. Furthermore, a close analysis of the reduced LD spectrum in the Soret band results in angles of approximately 80 degrees and 55 degrees between transition moments of the TMPyP and DNA helix axes in the absence of DAPI. All these observations indicate that the side of TMPyP whose structure resembles that of classical minor groove binding drugs does not fit deeply into the minor groove. This suggests that TMPyP binds across the minor groove: two positively charged pyridiniumyl rings interact electrostatically with negatively charged phosphate groups of DNA. When DAPI and TMPyP are simultaneously bound to poly(dA).poly(dT) or poly[d(A-T)(2)], the fluorescence intensity of DAPI decreases as TMPyP concentration increases, indicating that the excited energy of DAPI is transferred to TMPyP.

Binding properties of the tricationic porphyrin monomer with a phenolic substituent at the periphery and the porphyrin dimer conjugated with hydrophilic triethylene glycol were investigated in this study using absorption and polarized spectroscopy, namely, circular dichroism (CD) and linear dichroism (LD). The spectral properties of the porphyrin monomer, when complexed with polynucleotides, were essentially the same as that of the well-known meso-tetrakis(N-methylpyridiniumyl)porphyrin, indicating that the substitution at one peripheral pyridiniumyl ring did not affect the binding mode. When the porphyrin dimer formed a complex with poly[d(G-C)(2)], a negative CD band and a negative LD(r) spectrum were apparent in the Soret absorption region, with its LD(r) magnitude significantly smaller than that in the DNA absorption region. As the complex was stabilized over time, the intensity of the negative CD band and the negative LD(r) increased. These observations indicated that one of the porphyrin moieties of the dimer intercalated initially and than the other one also intercalated consecutively within a few hours. In the porphyrin dimer-poly[d(A-T)(2)] complex case, a bisignate CD was apparent and remained for at least 12 h, indicating that the porphyrins are stacked along the polynucleotide stem even at a very low [porphyrin]/[DNA base] ratio. A wavelength-dependent and time-dependent LD(r) of this complex suggests that the porphyrin molecular plane tilts strongly relative to the polynucleotide helix axis. The spectral properties of the porphyrin dimer-DNA complex are similar to those of the porphyrin dimer-poly[d(G-C)(2)] complex. However, some of the porphyrin moieties were located at the groove, which was evident by some positive characters in the CD and LD(r) spectra at the short wavelength in the Soret band.

Circular dichroism (CD) spectra of meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (TMPyP) that are associated with various duplex and triplex AT oligomers were investigated in this study. A strong positive CD was apparent for both the TMPyP complexed with duplex d[(A-T)(12)](2), d(A)(12).d(T)(12) and triplex d(A)(12).d[(T)(12)](2) at a low mixing ratio. As the mixing ratio increased, bisignate excitonic CD was produced for TMPyP complexed with duplexes, whereas the positive CD signal remained the same for the TMPyP-d(A)(12).d[(T)(12)](2) complex. This difference in the CD spectrum in the presence of duplex and triplex oligomers indicates that the moderate stacking of TMPyP occurs at the major groove of the duplex and the monomeric binding occurs in (or near) the minor groove. When TMPyP forms a complex with duplex d[(A-T)(6)](2) only excitonic CD was observed, even at a very low mixing ratio. Therefore, at least seven or more basepairs are required for TMPyP to exhibit a monomeric CD spectrum. After close analysis of the CD spectrum, the TMPyP-poly[d(A-T)(2)] complex could be explained by a combination of the CD spectrum of the monomeric, moderately stacked, and extensively stacked TMPyP.

Induced CD spectra of meso-tetrakis(N-methylpyridium-4-yl)porphyrin (TMPyP) complexed with d(A)12.d(T)12, d(G)12.d(C)12 duplex and d(A)12.[d(T)12]2, d(G)12.d(C)12.d(C)12+ triplex in the Soret band were compared in this study. When TMPyP is complexed with the duplex, a monomeric CD spectrum at a low [TMPyP]/[oligomer] ratio was apparent, while at a high mixing ratio, the excitonic CD was dominant. In contrast, when TMPyP was complexed with the triplex, the excitonic CD disappears at a relatively high mixing ratio, indicating the TMPyP exciton formation is inhibited by the third strand, which is located in the major groove. This observation indicates that the exciton is formed at the major groove of both AT- and GC-rich DNA, while the monomeric TMPyP binds at (or near) the minor groove of the AT site.

When meso-tetrakis(3-N-methylpyridiniumyl)porphyrin (m-TMPyP) formed a complex with poly[d(A-T)(2)], an intense bisignate excitonic CD in the Soret absorption region was observed. The excitonic CD of the m-TMPyP-poly[d(A-T)(2)] complex is unique in that no other combination of the related porphyrin, namely, meso-tetrakis(n-N-methylpyridiniumyl)porphyrin (where n = 2, 4), and polynucleotide including calf thymus DNA, poly[d(G-C)(2)], poly[d(I-C)(2)], and poly(dA).poly(dT), exhibits a comparable CD spectrum. From the [drug]/[DNA] ratio-dependence of the intensity and the shape of the CD spectrum, this porphyrin species is assigned to an extensively aggregated form. The extensively aggregated porphyrin disperses in 1 h after mixing to form moderately stacked porphyrin at a low mixing ratio. The magnitude of linear dichroism of the extensively aggregated porphyrin was small and the sign was negative in the Soret band, which indicated that the molecular plane of porphyrin in the complex is strongly tilted. On the other hand, the molecular plane of porphyrin is almost parallel to the DNA base plane (perpendicular to the DNA helix axis) in the moderately stacked form.

By utilizing circular and linear dichroism, the binding mode of meso-tetrakis(n-N-methylpyridiniumyl)porphyrin (n = 2, 3, 4) to various DNAs was studied in this work. 2-N-(methylpyridiniumyl)porphyrin(o-TMPyP), in which rotation of the periphery pyridinium ring is prevented, exhibits similar spectral properties when bound to DNA, poly[d(G-C)(2)] and poly[d(A-T)(2)], suggesting a similar binding mode. Close analysis of the spectral properties led us to conclude that o-TMPyP sits in the major groove. However, both 3-N- and 4-N-(methylpyridiniumyl)porphyrin (m- and p-TMPyP), of which the periphery pyridinium ring is free to rotate, intercalate between the basepairs of DNA and poly[d(G-C)(2)]. In the presence of poly[d(A-T)(2)], m-TMPyP exhibits a typical bisignate excitonic CD spectrum in the Soret band, while p-TMPyP shows two positive CD bands. The excitonic CD spectrum of the m-TMPyP-poly[d(A-T)(2)] complex and the positive CD band of the o-TMPyP-poly[d(A-T)(2)] complex were not affected by the presence of the minor groove binding drug, 4',6-diamidino-2-phenylindole (DAPI), indicating that this porphyrin is bound in the major groove. In contrast, two positive CD bands of the p-TMPyP-poly[d(A-T)(2)] complex altered in the presence of DAPI. From the changes in CD spectrum and other spectral properties, a few possible binding modes for p-TMPyP to poly[d(A-T)(2)] are suggested.

When meso-tetrakis(3-N-methylpyridiniumyl)porphyrin (m-TMPyP) formed a complex with poly[d(A-T)2], an intense bisignate excitonic CD in the Soret absorption region was observed. The excitonic CD of the m-TMPyP-poly[d(A-T)2] complex is unique in that no other combination of the related porphyrin, namely, meso-tetrakis(n-N-methylpyridiniumyl)porphyrin (where n = 2, 4), and polynucleotide including calf thymus DNA, poly[d(G-C)2], poly[d(I-C)2], and poly(dA)·poly(dT), exhibits a comparable CD spectrum. From the [drug]/[DNA] ratio–dependence of the intensity and the shape of the CD spectrum, this porphyrin species is assigned to an extensively aggregated form. The extensively aggregated porphyrin disperses in 1 h after mixing to form moderately stacked porphyrin at a low mixing ratio. The magnitude of linear dichroism of the extensively aggregated porphyrin was small and the sign was negative in the Soret band, which indicated that the molecular plane of porphyrin in the complex is strongly tilted. On the other hand, the molecular plane of porphyrin is almost parallel to the DNA base plane (perpendicular to the DNA helix axis) in the moderately stacked form. [ABSTRACT FROM PUBLISHER]

Examines the binding geometries of a series of Co3+-porphyrins and their free ligands using linear dichroism (LD), circular dichroism and fluorescence energy transfer. Negative LD of two non-metalloporphyrins; Cleavage rates of cobalt porphyrins.

In this study, the binding mode of porphyrin-free single-stranded poly[d(AT)] and trans-BMPyP was observed in the Z-form trans-BMPyP-poly[d(A-T)2] complex induced by extensive stacking depending on the temperature and concentration through circular dichroism (CD). The Z-form trans-BMPyP-poly[d(A-T)2] complex (R = 0.30) retained the Z-form DNA structure at a low temperature (20 °C) by the trans-BMPyP molecules. When the temperature was increased to 60 °C, the DNA was almost unfolded as a single-stranded poly[d(AT)], but the extensive stacking binding mode of trans-BMPyP was maintained and the shape of the porphyrin Soret band was symmetrically changed in comparison with the shape of the Z-form DNA. However, when the temperature was raised to 80 °C, the extensive stacking binding mode of trans-BMPyP was also unfolded almost completely. The binding mode of the trans-BMPyP-single-stranded poly[d(AT)] complex was very similar to the already known binding mode of porphyrins and a double-stranded DNA. The binding mode was dependent on the concentration ratio ([porphyrin]/[DNA]): a monomeric binding mode at a concentration ratio of 0.04, a moderate groove binding mode at a concentration ratio between 0.08 and 0.16, and extensive stacking at a concentration ratio between 0.20 and 0.30. The same result was obtained when the temperature of the Z-form DNA (R = 0.30) was increased to 60 °C. However, those binding modes were not found in cis-BMPyP, which was because, in the extensive stacking of trans-BMPyP along the DNA skeleton, the distance between the two positive methylpyridine ions at the trans site and thymine, one of the DNA bases, is decreased, creating a much more hydrophobic environment. In addition, the poly AT sequences found from the CD spectra for the binding of trans-BMPyP-poly[d(A-T)2] and trans-BMPyP-poly[d(AT)] (R = 0.30) showed that both of them underwent effective extensive stacking and that the chirality of extensive stacking was dependent on the form of DNA. [ABSTRACT FROM AUTHOR]

The effect of the number and position of the positive charges on porphyrin with respect to the mode of binding to poly[d(G-C)2] and poly[d(A-T)2] were investigated by absorption and polarized spectroscopy, including circular and linear dichroism (CD and LD). Meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (TMPyP), which possesses four positive charges on the periphery pyridinium rings, produces a negative CD and wavelength-independent reduced LD (LDr) spectra in the Soret absorption region when it associates with poly[d(G-C)2]. These spectral characteristics have been considered as diagnostic for intercalation. In contrast, both trans- and cis-bis(N-methylpyridinium-4-yl)diphenylporphyrin (trans- and cis-BMPyP), where the number of positive charges was reduced to two, multisignate CD and strong wavelength-dependence of the LDr spectra were observed, indicating that these porphyrins do not intercalate. Therefore, four positive charges are required for TMPyP intercalation. When associated with poly[d(A-T)2], trans-BMPyP exhibited a positive CD signal at a low [porphyrin]/[nucleobase] ratio with the appearance of a bisignate CD upon increase of the mixing ratio, suggestive of binding at the groove of the double helix at low mixing ratios, and stacking at increasing mixing ratios. Conversely, no monomeric binding was evident in the bis-BMPyP bisignate CD spectrum; hence, only the stacking mode was found for cis-BMPyP, even at the lowest [porphyrin]/[nucleobase] ratio, suggesting the importance of the position of the positive charges in determining monomeric groove binding or stacking. The binding geometries of trans- and cis-BMPyP were similar when associated with poly[d(A-T)2], as determined from the similar LDr spectrum. When associated with DNA, TMPyP exhibited similar spectral properties with that of the TMPyP−poly[d(G-C)2] complex, indicating intercalation of TMPyP between the DNA base pairs. Conversely, CD and LDr characteristics of both trans- and cis-BMPyP−DNA complexes resembled those that complexed with poly[d(A-T)2] at a high [porphyrin]/[DNA] ratio, suggesting that both porphyrins were stacked along the DNA stem. [ABSTRACT FROM AUTHOR]

Abstract: Interactions of water soluble cationic oxovanadyl (IV) meso-tetrakis(1-methylpyridium-4-yl)porphyrin (OVIV(TMPyP)4+) with double stranded poly[d(A-T)2], poly[d(G-C)2] nucleotides and calf thymus DNA were studied by the Raman spectroscopic techniques as well as the polarization spectroscopic measurements. The ground state Raman bands of OVIV(TMPyP)4+ observed in the presence of poly[d(A-T)2] were almost the same as those observed in a pure water solution except the slightly up-shifted Raman band (963cm−1) which is ascribed to the Ving mode of six-coordinated complex, OVIV(H2O)(TMPyP)4+. On the other hand, in the presence of poly[d(G-C)2], a markedly up-shifted Raman band was observed at 992cm−1, indicating that OVIV(TMPyP)4+ interacts with poly[d(G-C)2] by losing the sixth axial ligand. In the presence of calf thymus DNA, the Ving band was observed to be resulted by combination of those observed in poly[d(A-T)2] and poly[d(G-C)2]. The polarization spectroscopic studies with these results imply that OVIV(TMPyP)4+ interacts with DNA in different groove binding patterns originated from the formation of five-coordinated adducts in the G-C pair-rich regions and six-coordinated adducts in the A-T pair-rich regions. The down-shifts of core-size sensitive transient Raman bands (ν2 and ν4 modes) in the presence of nucleic acids illustrate an increase of the core-size of porphyrin macrocycle in the excited triplet state. The transient Raman bands related to pyridine group were observed, and they are interpreted to be due to the excited-state charge transfer from porphyrin ring to peripheral substituents. [Copyright &y& Elsevier]

Examines the binding of meso-Tetrakis(N-methylpyridium-4-yl)porphyrin to triplex oligonucleotides. Characteristics of the interaction between porphyrin and DNA; Effect of the third strand on the binding mode of the minor groove binding drugs and intercalators.