Light-scattering studies on deoxyribonucleic acid flexibility. The solution properties of a small circular deoxyribonucleic acid molecule

Previous investigations on the persistence length of DNA in solution have revealed large discrepancies between hydrodynamic results and those from light-scattering techniques which have potentially a greater resolving power. The information obtained from experiments on a small circular DNA molecule has resolved these discrepancies. The non-superhelical circular double-stranded DNA molecule from bacteriophage [unk]X174-infected cells is small enough to permit accurate light-scattering extrapolations, and its solutions have negligible anisotropy. The persistence length obtained from experimental investigations on this molecule is comparable with that obtained by hydrodynamic techniques, even with variation of the excluded-volume factor.     

Further studies on the deoxyribonucleic acid helping effect during transformation

Summary Transformation studies of the Challis strain of the H group of streptococcus were performed to further investigate the molecular basis of the deoxyribonucleic acid helping effect. Studies in the efficiency of transformation in the presence of non-transforming DNA support the notion that bacterial cells are indiscriminate in their uptake of donor DNA and that the helping effect occurs at a time when both transforming (T) and helping (H) DNAs have jointly entered the recipients. Furthermore, the ability of H DNA to promote transformation by T DNA is not directly altered by exposure of the former DNA to either UV-irradiation or nitrous acid. Nor does 5-bromouracil incorporation affect the capacity of H DNA to assist T DNA to transform a Challis cell.Increasing the concentration of denatured H DNA to a level that saturates the Challis bacteria in reaction mixtures produces a significant increase in the efficiency of genetic transformation. This increase in transformation frequency is greater with single DNA strands than with the corresponding amount of double strands.The extent of the helping effect with the Challis H DNA remains constant within an average molecular weight range of 3.5–7.0x10⁶ daltons. In the case of heterologous E. coli DNA, however, the helping function in this range is more pronounced as a result of decreasing molecular weight, even though the net incorporation of T DNA remains unaffected. When the average M. W. is reduced below 2×10⁶ a significant decline in the helping effect occurs in both cases.The effect of H DNA on the genetic transfer of two nonallelic antibiotic markers demonstrates that a saturating amount of non-transforming H DNA present in cells does not enhance the likelihood of co-integration of nonallelic factors. Evidence concerning the physiology of the competent cells and their ability to be helped reveals that the physiological basis of transformabilities and the helping capabilities of a culture are not identical.     

Kinetic and spectrophotometric studies on the renaturation of deoxyribonucleic acid

The kinetics of the renaturation of Escherichia coli DNA in 0.4-1.0m-sodium chloride at temperatures from 60 degrees to 90 degrees have been studied. The extent of renaturation was a maximum at 65 degrees to 75 degrees and increased with ionic strength, and the rate constant increased with both ionic strength and temperature. The energy and entropy of activation of renaturation were calculated to be 6-7kcal.mole(-1) and -40cal.deg.(-1)mole(-1) respectively. It has been shown that renaturation is a second-order process for 5hr. under most conditions. The results are consistent with a reaction in which the rate-controlling step is the diffusion together of two separated complementary DNA strands and the formation of a nucleus of base pairs between them. The kinetics of the renaturation of T7-phage DNA and Bordetella pertussis DNA have also been studied, and their rates of renaturation related quantitatively to the relative heterogeneity of the DNA samples. By analysis of the spectra of DNA at different stages during renaturation it was shown that initially the renatured DNA was rich in guanine-cytosine base pairs and non-random in base sequence, but that, as equilibrium was approached, the renatured DNA gradually resembled native DNA more closely. The rate constant for the renaturation of guanine-cytosine base pairs was slightly higher than for adenine-thymine base pairs.     

Sedimentation studies on the interaction of proflavine with deoxyribonucleic acid

A method is described of using photography to measure the concentrations of a small ligand (proflavine) above and below the boundary of a macromolecule (DNA, both native and denatured) sedimenting in the ultracentrifuge. The measurements are used to determine the extent of the binding of proflavine to DNA, and the results compared with those obtained by a spectrophotometric method. The results obtained by the two methods agree within 10%, thus validating the spectrophotometric technique under these conditions. The variation of the sedimentation coefficient with the extent of binding of proflavine was also studied. The results are discussed in relation to previously observed changes in the viscosity of the solutions.     

Denaturation mapping studies on the circular chloroplast deoxyribonucleic acid from pea leaves.

The structure of circular pea chloroplast DNA (ctDNA) has been analyzed by denaturation mapping. All of the pea ctDNA molecules that were examined had identical gross base sequences. Denaturation maps were constructed at denaturation levels of 2.5%, 22%, and 44%. These denaturation maps showed that the circular pea ctDNA contained six small AT-rich regions on one-half of the DNA molecule, and two small GC-rich regions on the other half of the DNA molecule. The structure of pea ctDNA circular dimers was also examined. The results showed that the pea ctDNA circular dimers consisted of two monomer length units integrated in tandem repeat.     

Spectroscopic and microcalorimetric studies on the molecular binding of food colorant acid red 27 with deoxyribonucleic acid

Interaction of the food colorant acid red 27 with double stranded DNA was investigated using spectroscopic and calorimetric methods. Absorbance and fluorescence studies suggested an intimate binding interaction between the dye and DNA. The quantum efficiency value testified an effective energy transfer from the DNA base pairs to the dye molecules. Minor groove displacement assay with Hoechst 33258 revealed that the binding occurs in the minor groove of DNA. Circular dichroism studies revealed that acid red 27 induces moderate conformational perturbations in DNA. Results of calorimetric studies suggested that the complexation process was driven largely by positive entropic contribution with a smaller favorable enthalpy contribution. The equilibrium constant of the binding was calculated to be (3.04 ± 0.09) × 10⁴ M^?1 at 298.15 K. Negative heat capacity value along with the enthalpy–entropy compensation phenomenon established the involvement of dominant hydrophobic forces in the binding process. Differential scanning calorimetry studies presented evidence for an increased thermal stability of DNA on binding of acid red 27. Copyright © 2016 John Wiley & Sons, Ltd.     

Quasi-elastic light scattering studies of hyaluronic acid solutions

Autocorrelation functions for the intensity of laser light scattered from solutions of hyaluronic acid have been measured under a variety of experimental conditions. The form of these functions is consistent with the large amount of long-range intra- and intermolecular interactions characteristic of polyelectrolyte solutions. Further experiments have been performed in order to study the effect of hyaluronic acid on the Brownian diffusion of polystyrene spheres. Using several different sphere sizes as solution probes, the importance of the ionic environment in determining hyaluronic confirmation has been demonstrated.