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Biomedical subjects

N Dattagupta

Publications and source records attributed to N Dattagupta.

At least 37 records · Page 2Linked to original sources
PubMed1981

Copper(II).bleomycin, iron(III).bleomycin, and copper(II).phleomycin: comparative study of deoxyribonucleic acid binding.

The kinetics and mechanism of binding of Cu-(II).bleomycin, Fe(III).bleomycin, and Cu(II).phleomycin to DNA were studied by using fluorometry, equilibrium dialysis, electric dichroism, and temperature-jump and stopped-flow spectrophotometry. The affinity of Cu(II).bleomycin for DNA was greater than that of metal-free bleomycin but less than that of Fe(III).bleomycin. Cu(II).bleomycin exhibited a two-step binding process, with the slow step indicating a lifetime of 0.1 s for the Cu(II).bleomycin.DNA complex. Fe(III).bleomycin binding kinetics indicated the presence of complexes having lifetimes of up to 22 s. DNA was lengthened by 4.6 A/molecule of bound Cu(II).bleomycin and by 3.2 A/bound Fe(III).bleomycin but not at all by Cu(II).phleomycin, suggesting that both bleomycin complexes intercalate while the phleomycin complex does not. However, phleomycin exhibited nearly the same specificity of DNA base release as bleomycin. These results suggest that the coordinated metal ion plays a major role in the binding of metal-bleomycin complexes to DNA but that intercalation is neither essential for DNA binding and degradation nor primarily responsible for the specificity of DNA base release by these drugs.

Animals↗
PubMed1981

Solution structural studies of the A and Z forms of DNA.

We report transient electric dichroism studies of short double-helical DNA and poly[d(G-C)] fragments in alcohol/water mixtures. The limiting reduced dichroism and the rotational correlation time changed abruptly in the alcohol concentration range expected for the DNA B-to-A and poly[d(G-C)] B-to-Z transitions. The Z form of poly[d(G-C)] was also induced by mitomycin C crosslinking in aqueous solution. The rotational correlation times observed for A- and Z-DNA were approximately consistent with dimensions determined by crystallographic and fiber diffraction analysis: the estimated rise per base pair was 2.8 A for A-DNA and 3.7 A for Z-DNA in solution. In addition, the observed limiting reduced dichroism values for A- and Z-DNA were close to the theoretical limit of -1.5, requiring a structure in which the base transition moments are effectively perpendicular to the double-helix axis. This is the result expected for any DNA double helix having dyad symmetry in which the base pairs are flat and the base transition moments lie predominantly in the short axis of the base pair and therefore close to a helix dyad axis. Only B-DNA deviates from this rule, strongly reinforcing our earlier conclusion that the base pairs in B-DNA are not flat but are propeller twisted, either statically or as a dynamic average.

Animals↗
PubMed1981

Conversion of B DNA between solution and fiber conformations.

At low salt and high DNA concentration, rodlike DNA molecules form specific aggregates containing about seven parallel DNA molecules packed in a bundle. Bundle formation can be reversed by addition of small amounts of EDTA, and formation of the specific aggregates can be induced at higher salt concentration by addition of trivalent ions. In contrast to monomeric DNA, the aggregated species has the optical anisotropy expected for the B form as observed in fibers.

Animals↗
PubMed1980

Interaction of netropsin and distamycin with deoxyribonucleic acid: electric dichroism study.

We report dichroism and equilibrium binding studies of netropsin (Net) and distamycin A3 (Dist) binding to deoxyribonucleic acid (DNA). We show that at low degrees of binding (r) to calf thymus DNA, Net induces a considerable increase in the apparent DNA length (14 A/drug molecule bound), closely analogous to the results reported earlier for Dist. In addition, we show that chicken erythrocyte DNA shows length changes similar to those of calf thymus DNA upon distamycin binding. DNA length reaches a maximum at 1 bound drug/20-30 base pairs and then decreases to its initial value by r = 0.1. This effect is not seen for two other DNAs with nearly identical A + T base pair content and may therefore arise from the details of base sequence or base modification in eukaryotic DNA. We also show that Dist binding to calf thymus DNA at low r values is positively cooperative and shows a DNA affinity which is primarily nonionic. We demonstrate that independent of the DNA to which they are bound, the Net and Dist transition moments are inclined by 43 +/- 3 degrees from the helix axis, consistent with the idea that both drugs bind inside and parallel to the DNA small groove. From dichroism measurements, we show that the conformational change induced in calf thymus DNA by Dist does not kink or bend the helix and does not substantially alter the average inclination of the bases. Finally, we outline a statistical mechanical theory for calculation of binding isotherms when binding is coupled to a DNA structural change.

Animals↗
PubMed1980

Unfolding of nucleosomes by ethidium binding.

We report spectroscopic, hydrodynamic, and biochemical studies on the complex of ethidium bromide with 140 base pair nucleosomal core particles. Fluorescence titration indicates a greater intrinsic affinity of ethidium for nucleosomes than for DNA, and fluorescence depolarization measurements imply increased immobilization of ethidium bound to nucleosomes, but with more extensive dye-dye energy transfer compared to DNA-bound dye. Ethidium intercalated into DNA in nucleosomes has a limiting reduced linear dichroism of -0.45 at 320 nm and -0.25 at 530 nm. Both the energy transfer and dichroism results are consistent with clustering of the nucleosome-bound dye molecules. Electric dichroism measurements and ultracentrifugation studies reveal that structural distortion of the nucleosome accompanies ethidium binding, occurring in the range of r (ethidium residues per base pair) values from 0.02 to 0.06. The distortion transition is characterized by an increase in the negative limiting reduced dichroism from 0.29 to 0.45 at 265 nm, an increase in the field-induced viscosity-limited rotational orientation time from 0.8 to 3 mus, and a decrease in sedimentation coefficient from 10.5 to 8.2 S. The complex was modeled hydrodynamically as a cylinder of 335-A length and 67-A diameter, containing 1.4 superhelical turns of DNA. Dimethylsuberimidate cross-linked nucleosomes, or native nucleosomes in the presence of Mg2+, bind ethidium weakly and are not distorted. The periodicity of cutting sites produced by DNase II digestion of nucleosomes remains constant as ethidium is added, but the bandwidth increases. A thermodynamic model is proposed to interpret the binding isotherm, based on enhancement of drug binding affinity due to release of superhelical stress in the nucleosome-ethidium complex.

Base Composition↗
PubMed1979

Selective repression of transcription by base sequence specific synthetic polymers.

We report the effect of novel synthetic polymers on deoxyribonucleic acid (DNA) directed ribonucleic acid (RNA) synthesis in vitro. Polymers contained base-selective monomers, including a GC-specific phenazine derivative and an AT-specific triphenylmethane dye. Radical chain polymerization was carried out in aqueous solution by using monomers bound to a template DNA, which was obtained from either lambda or T7 bacteriophage. Polymers were isolated and reannealed with DNA samples, including competitive mixtures of T7 and lambda DNAs. We measured transcription from DNA-polymer complexes by using Escherichia coli RNA polymerase and determined not only the reduction in total transcription levels but also the relative inhibition of lambda- or T7-specific transcription by using a hybridization assay. The results show that micromolar concentrations of individual dyes are sufficient to cause substantial inhibition of transcription when the dyes are incorporated into polymers. More significantly, a number of the polymers inhibited more strongly transcription from the DNA which had served as template for polymer synthesis than from the DNA present as competitor in the annealing process. We conclude that template synthesis of DNA-binding polymers can lead to preferential inhibition of function of the original template. The apparent relative affinity of polymer for competing DNAs can be altered by at least an order of magnitude depending on which DNA was used as the synthesis template. The results offer a new approach to improving the specificity of DNA-binding drugs.

Coloring Agents↗
PubMed1979

Transmission of allosteric effects in DNA.

Binding of distamycin induces a cooperative transition of calf thymus DNA to a new form with higher affinity for the drug and altered structural properties.

Allosteric Regulation↗
PubMed1979

Binding of bleomycin to DNA: intercalation of the bithiazole rings.

At pH 5.5, binding of bleomycin relaxed supercoiled ColE1 DNA without breaking it. Binding of tripeptide S, a fragment of the drug containing the bithiazole rings, also relaxed and then recoiled supercoiled DNA, at pH 5.5 and at pH 8.0, where bleomycin is normally active. The unwinding angle was 12 degrees. Both compounds lengthened linear DNA by 3.1 A per molecule bound, and linear dichroism (303--315 nm) of bleomycin bound to linear DNA oriented in an electric field indicated the presence of a chromophore making an angle of 59--61 degrees with the helix axis. These results strongly suggest that bleomycin binding to DNA involves intercalation of the bithiazole rings. In 0.1 M Na Cl at pH 8, supercoiled ColE1 DNA was broken at a rate 50% greater than relaxed closed circular ColE1 DNA. Since supercoiling increases the affinity of DNA for intercalators, this result suggests that intercalative binding is involved in bleomycin-induced breakage of DNA.

Bleomycin↗
PubMed1979

Structure of DNA within three isometric bacteriophages.

This paper describes a model for the structure of DNA contained in three morphologically similar bacteriophages--T7, P22 and phiCd-1--based on the transient electric dichroism of intact phage. The reduced dichroism of each of the phages at perfect orientation is within the range +0.12 to +0.19. Assuming that the phage orientation axis is that which passes from the apex through the tail, the measured dichroism suggests that DNA is wrapped in closely packed, co-axial solenoids with the axis of the solenoids tipped 43.5 degrees +/- 2.5 degrees from the orientation axis of the phage. All three phages show a large permanent dipole moment, with respective values of 5600, 200,000 and 500,000 Debye for T7, phiCd-1 and P22. The radius of the equivalent sphere for the three phages calculated from the rotational relaxation time for the rise of dichroism is in agreement with birefringence and electron microscope observations. The circular dichroism spectra of all three bacteriophages indicate that the local DNA helicity is similar in each case.

Coliphages↗
PubMed1978

Transient electric dichroism studies of nucleosomal particles.

We report transient electric dichroism experiments on nucleosomal core particles containing 140 and 175 base pairs of DNA, and on spacerless dinucleosomes. The results indicate that all particles posses a permanent dipole moment. The orientation time of 140 base pair nucleosomes implies an estimated maximum dimension of a = 130 A (a must be at least 111 A), consistent with the disk model. The maximum dimension of the spacerless dinucleosome is estimated to be about 290 A (at least 180 A), ruling out a structure in which two disks are stacked directly on top of each other. The reduced dichroism amplitude indicate that the DNA superhelix axis in nucleosomes aligns perpendicular to the electric field, as expected for a dipole moment directed along a C2 symmetry axis across the disk diameter. Nucleosomes containing 175 base pairs of DNA show a substantially larger dichroism amplitude that do 140 base pair nucleosomes. In the context of the disk model, this result is shown to be consistent with 100 base pairs of DNA per superhelical turn, but not with 80 base pairs per turn.

Animals↗
PubMed1978

Physical studies of nucleosome assemble.

Biochemical, spectroscopic, and hydrodynamic studies were performed on the reconstituted complex of 140 base pair DNA and the arginine-rich histone tetramer (H3/H4)2. The histones bind to DNA in a 1:1 molar ration to form a stable particle which orients in an electric field with a rotational correlation time of 6.3 mus and a limitign reduced dichroism of --0.74. The complex was modeled hydrodynamically as a cylinder of dimensions 450 X 80 X 80 A containing approximately 1.5 superhelical turns. Addition of the lysinerich histones to this complex cause a condensation of the structure and results in physical properties nearly identical with those of a native nucleosomal particle.

Animals↗
PubMed1978

Transient electric dichroism of rod-like DNA molecules.

We report transient electric dichroism studies on monodisperse rod-like DNA molecules. By using restriction fragments and DNAs of known length, it is shown that the orientation time is accurately predicted by the theoretically calculated rotational diffusion coefficient. The field dependence of the steady-state dichroism values is not consistent with the induced electric dipole orientation mechanism, and the time dependence is not consistent with the presence of a permanent dipole moment. In order to explain the dependence of the dichroism on the electric field, the ionic strength of the medium, and the length of the macromolecule, we propose a new model in which anisotropic ion flow produces an asymmetric ion atmosphere around the polyelectrolyte, resulting in an orienting torque. From the limiting dichroism at high field, we estimate that the DNA bases are inclined at an angle of 73 degrees or less relative to the helix axis, in good agreement with the revised model of B-form DNA suggested by Levitt, in which the base pairs have a propeller-like twist. Our results establish transient electric dichroism measurements as a technique well suited for study of alterations in the length and base pair inclination of rod-like DNA molecules.

Anions↗
PubMed1978

Does irehdiamine kink DNA?

We report equilibrium, relaxation kinetic, and transient electric dichroism studies on the complex of the diamino steroid irehdiamine A with DNA. The results are consistent with a beta-kinked structure for the complex at saturation, with a kink in the DNA structure induced by a bound steroid every second base pair. The results that favor this hypothesis include an apparent length decrease of rod-like bacterial DNA molecules when only a small amount of drug is bound, followed by an apparent length increase at saturation. The limiting dichroism amplitude implies a substantial increase in the tilt of the bases relative to the orientation axis; at saturation the base UV transition moments are tilted about 31 degrees from the plane perpendicular to the orientation axis. Because of the direction of polarization of the 260-nm transition moments, the results indicate that the tilt of the bases must be predominantly in the short rather than the long axis of the base pair. The large hyperchroism of the complex is consistent with loss of base stacking, as required by a kinked structure. The kinetic results imply a bimolecular reaction mechanism, with a temperature-dependent association rate constant of roughly 10(8) M(-1) sec(-1), and a dissociation rate constant of about 5 x 10(3) sec(-1), nearly independent of temperature. The association activation energy and apparent reaction enthalpy vary from 12 to 22 kcal mol(-1); heat is absorbed on complex formation as expected for loss of base-stacking interactions. An anomalous result of the experiments is the larger apparent length increase (13%) exhibited by two eukaryotic DNAs, compared to 6% for three prokaryotic DNAs. Differences were also observed in the kinetic properties of the complexes.

Binding Sites↗
PubMed1975

Interaction of 4,5-dibromo-2,7-di-(acetatomercuri)-fluorescein with DNAs of different base composition.

The changes in absorption spectra in the visible region observed on adding different naturally occurring and synthetic DNA duplexes to solutions of 4,5-dibromo-2,7-di-(acetatomercuri)-fluorescein indicate that the mercurial reacts with polynucleotides of this type. The reaction is reversible as proved by adding excess of KCN which restores the original spectra of the free dye. The interaction is characterised also by quenching of the fluorescence of the dye and the induction of optical activity in it. The extent of these spectral effects depends strongly on the (A+T) content of the complexed DNA and decreased in the order: poly [d(A-T)], Clostridium perfringens DAN, Escherichia coli DNA, Micrococcus luteus DNA and poly(dC). From equilibrium-dialysis experiments the same order in affinity is obtained when these poly-nucleotides are at equilibrium with the same concentration of 4,5-dibromo-2,7-di-(acetatomercuri)-fluorescein. From the changes produced by different mercurials in the ORD spectra and viscosity of a DNA solution it has been concluded that 4,5-dibromo-2,7-di(acetatomercuri)-fluorescein does not cause any drastic alteration of the secondary structure of DNA.

Animals↗
PubMed1975

Interactions of heteroaromatic compounds with nucleic acids. 2. Influence of substituents on the base and sequence specificity of intercalating ligands.

This paper presents the results of a systematic study on the effects of substituents on the base and sequence specificity of tricyclic heteroaromatic compounds interacting with DNA by intercalation. All the compounds tested are derived from proflavine and acridine orange analogs with different heteroatoms in the middle ring. Their base and sequence specificities were determined by differential dialysis of the ligand against DNA samples of differing G-C content. The main results indicate that (a) the introduction of a phenyl substituent into one of the two available positions of the middle ring increases or decreases the G-C specificity of the ligand depending on the position where the substitution takes place; (b) compounds of the substitution type of neutral red (2-methyl-3-amino-7-dimethyl-amino-phenazine) show unexpectedly high G-C specificities and (c) DNA ligands of pronounced sequence specificity for adjacent G-C pairs can be constructed by combining the structural elements of neutral red with an additional phenyl residue in the same molecule. The further study of compounds related to the phenylated neutral red revealed that the G-C specificity can be improved or destroyed by additional substituents. The comparison of the G-C specificity and the DNA-affinity data of the compounds studied leads to the suggestion that the specificity arises mainly from electronic factors which are strongly controlled through steric constraints on possible ocmplex geometries. As a basis for the discussion a possible structure for the DNA complex of the phenylated neutral red is considered in which the extra phenyl ring at N-5 of the phenazinium system, protrudes into the large groove of the DNA helix while the tricyclic part of the ligand is inserted between the DNA base-pairs.

Acridines↗