DNA proflavine

Fig. 11 Polarized absorption spectra of DNA /proflavine/2Ci8-glu-N LB films (88 layers) deposited by a vertical dipping method and b horizontal lifting method...

Kinetics. In the observed relaxation curves, there are two well separated relaxation times, usually differing about tenfold in time scale. In the native DNA-proflavine system, however, the amplitude for fast process was so small that the faster relaxation time could not be measured with high precision. 

The DNA-proflavine complex exhibits an X-ray diffraction pattern which is characteristically modified. 

A good understanding of the properties of water is thus essential as we move to more complicated systems. We have been involving in the study of aqueous solution of many important biological molecules, such as acetylcholine, Gramicidin, deoxydinucleoside phosphate and proflavin, and DNA, etc., first at the Monte Carlo level and slowly moving to the molecular dynamics simulations. We will discuss some of the new results on the hydration structure and the dynamics of B- and Z-DNA in the presence of counterions in the following. 

As described in section 4.1, the DNA double helix must unwind to allow access ofthe polymerase enzymes to produce two new strands ofDNA. This is facilitated by DNA gyrase, the target of the quinolones. Some agents interfere with the unwinding of the chromosome by physical obstruction. These include the acridine dyes, of which the topical antiseptic proflavine is the most familiar, and the antimalarial acridine, mepacrine. They prevent strand separation by insertion (intercalation) between base pairs from each strand, but exhibit very poor selective toxicity. 

Koglin E., Sequaris J.M., Interaction of proflavine with DNA studied by colloid surface enhanced resonance Raman-spectroscopy, J. Molecular Struct. 1986 141 405-409. 

Thus complete intercalation of the aromatic PAH between the bases of DNA, in the manner described above for flat molecule such as proflavine, did not seem to be a likely mechanism for the carcinogenic action of these compounds. Since alkylation and intercalation are not simultaneously possible for steric reasons, and since one molecule is wedge-shaped and the other is flatter, it was considered more likely that the action of these compounds arose from their alkylating ability they could alkylate a base of DNA and then, since the bulky aromatic hydrophobic group would possibly not remain protruding into the hydrophilic environment, it is possible that the aromatic PAH group could then lie in one of the grooves of DNA. 

Acridine derivatives, such a proflavine (5) and acridine orange (6) (Scheme 3), are a second class of intercalative DNA guest molecules for which binding dynamics have been extensively studied. 

Table 3 Rate constants for the binding of proflavine (5) to DNA assuming the mechanism shown in Scheme 4.

Potentiometric titrations, in alcohol, 276-278 with metal ions, 278-284 solvation effects, 277-278 Proflavine, binding with DNA, 190-194 re-systems, orthogonal, 3 in-plane and out-of-plane, 7-10... 

A related observation is that fully relaxed supercoiled DNA/dye complexes are somehow different from nicked circular DNA/dye complexes in the presence of the same concentration of free dye, where the binding ratios should be the same. This is readily seen in gel electrophoresis in the presence of sufficient dye concentration so that at least one, but not all, of the topoisomers is positively supercoiled. The slowest moving, presumably fully relaxed, topoisomer migrates significantly faster than the nicked circle, and this difference increases with the amount of dye present. This is not observed with chloroquine, perhaps because the effect is too small. However, it is readily apparent in the original gels of Keller0 61) in which ethidium was used to unwind the topoisomers. We have confirmed this effect for ethidium and have observed similar behavior for proflavine, 9-aminoacridine, and quinacrine. 

The steady-state FPA of large ( 500bp) calf thymus DNA/ethidium complexes is unaffected by addition of proflavine up to one per two base pairs. From this, it is concluded that the torsion constant is unaltered(168) by intercalation of proflavine. However, in our time-resolved FPA studies of linear pBR322 DNA/ethidium complexes, the torsion constant is reduced by the factor 0.60 as proflavine is added from zero to one per two base pairs.(173) Whether this discrepancy is due to a real difference between these DNAs or... 

When the stretched DNA-lipid film was soaked in an aqueous solution of ethidium bromide (itmax = 480 nm) for a day at room temperature, the transparent film turned red (itmax = 520 nm) and the aqueous solution became clear (Fig. 9a). Thus, the ethidium intercalated completely between base pairs of the DNA film. When the film was moved into the new aqueous buffer solution, the intercalated dye molecules were hardly removed from the film at least for a day. Similar intercalation behavior into the film was observed for other dyes such as proflavine, acridine orange, and safranine T [14-17]. 

Cationic amphiphiles 2Ci8-glu-N spread on pure water, in the solution of 10 xM DNA containing 10 xM intercalating dyes (proflavine). The dye-intercalated DNA anions were expected to adsorb to the cationic lipid mono-layer due to electrostatic interactions and was transferred to a hydrophobized glass plate at a surface pressure of 35 mN m at 20 °C. From a moving area of a barrier, two layers of the monolayer were confirmed to be transferred in each one cycle (Y-type deposition). When the QCM plate was employed as a transfer plate, the transferred mass could be calculated from frequency decreases (mass increase on the QCM) [29-31]. It was confirmed that 203 10 ng of two lipid monolayers and 74 5 ng of DNA strands were transferred on to the substrate per dipping cycle, which means ca. 95% of the monolayer area was covered by DNA molecules. 

The presence of proflavine protects some viruses against inactivation by ultraviolet radiation and proflavine has been shown to inhibit formation of thymine dimers in DNA.86 (6) The sensitivity of microorganisms to ultraviolet light increases with, but is not proportional to the square of, the thymine content of the DNA.86 (7) Substitution of... 

Figure 5.13 shows the structures of extrinsic fluors that have been of value in studying biochemical systems. ANS, dansyl chloride, and fluorescein are used for protein studies, whereas ethidium, proflavine, and various acridines are useful for nucleic acid characterization. Ethidium bromide has the unique characteristic of enhanced fluorescence when bound to double-stranded DNA but not to single-stranded DNA. Aminomethyl coumarin (AMC) is of value as a fluorogenic leaving group in measuring peptidase activity. 

Mauss Y, Poulet P, Steibel J, et al. 1980. Interaction of proflavine-calf thymus DNA complexes with Ag+ and Hg++ ions. Studia Biophysica 81 95-96. 

We first describe the NMR parameters for the duplex to strand transition of the synthetic DNA poly(dA-dT) (18) with occasional reference to poly(dA-dU) (24) and poly(dA- brdU) and the corresponding synthetic RNA poly(A-U) (24). This is followed by a comparison of the NMR parameters of the synthetic DNA in the presence of 1 M Na ion and 1 M tetramethylammonium ion in an attempt to investigate the effect of counterion on the conformation and stability of DNA. We next outline structural and dynamical aspects of the complexes of poly(dA-dT) with the mutagen proflavine (25) and the anti-tumor agent daunomycin (26) which intercalate between base pairs and the peptide antibiotic netropsin (27) which binds in the groove of DNA. 

Planar dyes such as proflavine can bind to DNA by intercalation between base pairs or by stacking along the groove of the helix. The former process is favored when the nucleic acid is in excess (Nuc/D 4) and in high salt solution. We are interested in the intercalation process and hence our NMR studies have focussed on an investigation of the proflavine poly(dA-dT) complex as a function of the Nuc/D ratio in 1 M NaCl in an attempt to probe the structure and dynamics of mutagen-nucleic acid interactions in solution (25). 

Nuc/D =8, in 1 M NaCl solution with the resonance shifting to high field on complex formation (Figure 16). The results demonstrate that the base pairs are intact in the proflavine complex with the synthetic DNA. 

Melting Transition Typical 360 MHz proton NMR spectra of proflavine poly(dA-dT) complexes, Nuc/D = 24 and Nuc/D = 8, in 1 M NaCl solution at temperatures below the midpoint for the dissociation of the complex are presented in Figures 17A and B respectively. The stronger base and sugar resonances can be readily resolved from the weaker proflavine resonances (designated by asterisks) in the presence of excess nucleic acid (Figure 17) so that the resonances of the synthetic DNA and the mutagen can be monitored independently of each other. 

Overlap Geometry A schematic representation of the proposed overlap geometry for proflavine intercalated into a deoxy pyrimidine(3 -5 )purine site is presented below with the (o) symbols representing the location of the phenanthridine ring protons. The mutual overlap of the two base pairs at the intercalation site involves features observed in the crystal structures of a platinum metallointercalator miniature dC-dG duplex complex (55) and the more recent proflavine miniature dC-dG duplex complex (48), as well as features derived in a linked-atom conformational calculation of the intercalation site in the proflavine DNA complex (51). [4]... 

The overlap of proflavine with adjacent base pairs was varied until there was approximate agreement between the experimental upfield complexation shifts (Table V) and those calculated from ring current and atomic diamagnetic anisotropy contributions from the base pairs (56). The calculated upfield shifts are somewhat smaller than the experimental complexation shifts at the proflavine protons in the synthetic DNA complex (Table V). This... 

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