Characterization of the binding of paylean and DNA by fluorescence,UV spectroscopy and molecular docking techniques

The interaction of paylean (PL) with calf thymus DNA (ctDNA) was investigated using fluorescence spectroscopy, UV absorption, melting studies, ionic strength, viscosity experiments and molecular docking under simulated physiological conditions. Values for the binding constant K_a between PL and DNA were 5.11 × 10³, 2.74 × 10³ and 1.74 × 10³ L mol^–1 at 19, 29 and 39°C respectively. DNA quenched the intrinsic fluorescence of PL via a static quenching procedure as shown from Stern–Volmer plots. The relative viscosity and the melting temperature of DNA were basically unchanged in the presence of PL. The fluorescence intensity of PL–DNA decreased with increasing ionic strength. The value of K_a for PL with double‐stranded DNA (dsDNA) was larger than that for PL with single‐stranded DNA (ssDNA). All the results revealed that the binding mode was groove binding, and molecular docking further indicated that PL was preferentially bonded to A–T‐rich regions of DNA. The values for ΔH, ΔS and ΔG suggested that van der Waals forces or hydrogen bonding might be the main acting forces between PL and DNA. The binding distance was determined to be 3.37 nm based on the theory of Förster energy transference, which indicated that a non‐radiation energy transfer process occurred. Copyright © 2015 John Wiley & Sons, Ltd.     

Thermodynamic and structural study of phenanthroline derivative ruthenium complex/DNA interactions: probing partial intercalation and binding properties

The binding of [Ru(PDTA-H₂)(phen)]Cl (PDTA = propylene-1,2-diaminetetra-acetic acid; phen = 1,10 phenanthroline) with ctDNA (=calf thymus DNA) has been investigated through intrinsic and induced circular dichroism, UV-visible absorption and fluorescence spectroscopies, steady-state fluorescence, thermal denaturation technique, viscosity and electrochemical measurements. The latter indicate that the cathodic and anodic peak potentials of the ruthenium complex shift to more positive values on increasing the DNA concentration, this behavior being a direct consequence of the interaction of both the reduced and oxidized form with DNA binding. From spectrophotometric titration experiments, the equilibrium binding constant and the number of monomer units of the polymer involved in the binding of one ruthenium molecule (site size) have been quantified. The intrinsic circular dichroism (CD) spectra show an unwinding and a conformational change of the DNA helix upon interaction of the ruthenium complex. Quenching process, thermal denaturation experiments and induced circular dichroism (ICD) are consistent with a partial intercalative binding mode.     

Synthesis of novel bispyrene diamines and their application as ratiometric fluorescent probes for detection of DNA

Fluorescent DNA probes with 1,6-hexanediyl as the linker between two pyrenes, phenylpyrenes or phenylethynyl pyrene fluorophores were synthesized (Py-1, Py-2 and Py-3) and their interactions with DNA were studied by UV–vis absorption spectra, fluorescence spectra and viscosity measurements. The probes show red-shifted emission compared with pyrene (up to 20 nm). We found the interaction of these probes with DNA can be either intercalation or groove binding. Ratiometric fluorometry (ratio of the monomer and excimer emission intensity versus concentration of DNA) was achieved with these probes for DNA quantification (with limit of detection, LOD, up to 0.1 μg/mL). We also found that the undesired oxygen sensitivity of the emission intensity of pyrene fluorophore can be greatly suppressed by extending the π-conjugation framework of pyrene (the I_Ar/I_air value is decreased from 8.10 for pyrene to less than 2.20 for the DNA probes described herein).     

The structure of the chloroplast genome in members of the genus Asparagus

 In a previous study we constructed a physical map of the chloroplast DNA (ctDNA) of garden asparagus (Asparagus officinalis L. cv ‘Mary Washington 500W’; Lee et al. 1996). In the present study we have constructed and compared HindIII and XhoI restriction maps of the ctDNAs of eight species of Asparagus: namely, A. officinalis, A. schoberioides, A. cochinchinensis, A. plumosus, A. falcatus, A. sprengeri, A. virgatus and A. asparagoides. The ctDNA of A. officinalis has 32 and 23 sites that are recognized by HindIII and XhoI, respectively. Taking the physical map of the ctDNA of A. officinalis as a standard, we found that the ctDNAs of A. falcatus, A. sprengeri, and A. asparagoides each had one additional HindIII site and lacked one XhoI site. We also detected two relatively large deletions of nucleotides in the ctDNA from A. cochinchinensis by sequencing analysis. Both of these deletions were located in a non-coding region between the ndhC and trnV genes and were 95 bp and 347 bp in length, respectively. The regions around the deletions exhibited strong homology, and short direct-repeat sequences were detected at the borders of the deletions, an indication that these deletions were the result of intramolecular recombination mediated by the direct repeats. Received: 16 June 1997 / Accepted: 17 July 1997     

Genetic profiling of cancer with circulating tumor DNA analysis

Circulating cell-free tumor DNA(ctDNA) in the blood is DNA released from apoptotic, circulating, and living tumor cells. ctDNA is about 140 nt in length and has a half-life of about 1.5 h. ctDNA analysis provides a noninvasive means to assess the genetic profile of cancer in real time. With the advent of molecular technologies, including digital PCR and massively parallel sequencing(MPS), ctDNA analysis has shown promise as a highly sensitive and specific alternative to conventional tissue biopsy in cancer detection, longitudinal monitoring, and precision therapy. This review provides an overview of the latest development in our understanding of the biologic characteristics, detection methodologies, and potential clinical implications of ctDNA, as well as the challenges in translating ctDNA analysis from the research arena to patient care.     

The physical map of the chloroplast DNA from Asparagus officinalis L.

The genus Asparagus consists of 100–300 species of both dioecious and hermaphrodite plants. Since there are diploid, tetraploid, and hexaploid plants in this genus, RFLP (restriction fragment length polymorphism) analysis of chloroplast DNA (ctDNA) is suitable for examining the phylogenetic relationships. We have constructed a physical map of the ctDNA of garden asparagus (A. officinalis L. cv Mary Washington 500 W) using five restriction endonucleases, namely, BamHI, PstI, SalI, HindIII, and XhoI. Asparagus ctDNA was digested with restriction enzymes and cloned into plasmid and phage vectors, and a clone bank was constructed that covered 70% of the genome. A physical map was constructed by Southern hybridization of total DNA from asparagus with homologous and heterologous probes. The asparagus ctDNA was about 155 kb long and it contained two inverted repeats (23kb each) separated by a large single-copy region (90kb) and a small single-copy region (19kb). Fifteen genes, encoding photosynthesis-related proteins, rDNAs, and tRNAs, were localized on the physical map of asparagus ctDNA. Comparing the length and the gene order of asparagus ctDNA with that of other plants, we found that asparagus ctDNA was similar to tobacco ctDNA but different from rice ctDNA. The restriction patterns of the ctDNAs from several varieties of A. officinalis and three species of Asparagus were analyzed. The restriction patterns of the varieties of A. officinalis were very similar, but polymorphisms were detected among the three species of Asparagus.     

Biparental inheritance of chloroplast DNA and the existence of heteroplasmic cells in alfalfa

Summary Mapping of chloroplast DNA (ctDNA) restriction fragment patterns from a chlorophyll deficient mutant and two phenotypically normal alfalfa genotypes (Medicago sativa L.) has demonstrated the existence of a distinct ctDNA genotype from each source. These unique restriction fragment patterns were utilized to identify maternal or paternal origin of ctDNA in hybrid plants from crosses involving the normal alfalfa genotypes as females and the yellow-green chlorophyll deficient sectors as males. Progeny from these crosses expressing the yellow-green sectored phenotypes contained paternal ctDNA in the chlorophyll deficient sectors and maternal ctDNA in the normal sectors, confirming biparental plastid inheritance. The existence of mixed cells containing both mutant and normal plastids at various stages of sorting-out was observed by transmission electron microscopy of mesophyll cells in mosaic tissue from hybrid plants. This observation verified the biparental transmission of plastids in alfalfa.     

Interspecific somatic hybridization between Brassica napus and Brassica hirta (Sinapis alba L.)

Summary Somatic hybridization between Brassica napus and B. hirta (or Sinapis alba) is described. No cybrid plant with B. napus nucleus exhibiting cytoplasmic male sterility was recovered. Somatic hybrids were identified morphologically and, for some of them, by cytological observations. They were also characterised by Southern hybridization of nuclear rDNA. Chloroplast and mitochondrial DNA restriction analysis showed that 2 plants out of 14 have B. hirta ctDNA, one the B. napus mtDNA and the other a hybrid. Nine possess B. napus ctDNA with a hybrid mtDNA. For six of them, mtDNA patterns present novel bands, suggesting intergenomic recombination during fusion. These hybrids will be included in the breeding program.     

Saffron and natural carotenoids: Biochemical activities and anti-tumor effects

Saffron, a spice derived from the flower of Crocus sativus, is rich in carotenoids. Two main natural carotenoids of saffron, crocin and crocetin, are responsible for its color. Preclinical studies have shown that dietary intake of some carotenoids have potent anti-tumor effects both in vitro and in vivo, suggesting their potential preventive and/or therapeutic roles in several tissues. The reports represent that the use of carotenoids without the potential for conversion to vitamin A may provide further protection and avoid toxicity. The mechanisms underlying cancer chemo-preventive activities of carotenoids include modulation of carcinogen metabolism, regulation of cell growth and cell cycle progression, inhibition of cell proliferation, anti-oxidant activity, immune modulation, enhancement of cell differentiation, stimulation of cell-to-cell gap junction communication, apoptosis and retinoid-dependent signaling. Taken together, different hypotheses for the antitumor actions of saffron and its components have been proposed such as a) the inhibitory effect on cellular DNA and RNA synthesis, but not on protein synthesis; b) the inhibitory effect on free radical chain reactions; c) the metabolic conversion of naturally occurring carotenoids to retinoids; d) the interaction of carotenoids with topoisomerase II, an enzyme involved in cellular DNA-protein interaction. Furthermore, the immunomodulatory activity of saffron was studied on driving toward Th1 and Th2 limbs of the immune system. In this mini-review, we briefly describe biochemical and immunological activities and chemo-preventive properties of saffron and natural carotenoids as an anticancer drug.     

Molecular interaction of ctDNA and HSA with sulfadiazine sodium by multispectroscopic methods and molecular modeling

Interactions of sulfadiazine sodium (SD‐Na) with calf thymus DNA (ctDNA) and human serum albumin (HSA) were studied using fluorescence spectroscopy, UV absorption spectroscopy and molecular modeling. The fluorescence experiments showed that the processes were static quenching. The results of UV spectra and molecular modeling of the interaction between SD‐Na and ctDNA indicated that the binding mode might be groove binding. In addition, the interaction of SD‐Na with HSA under simulative physiological conditions was also investigated. The binding constants (K) and the number of binding sites (n) at different temperatures (292, 302, 312 K) were 5.23 × 10³ L/mol, 2.18; 4.50 × 10³ L/mol, 2.35; and 4.08 × 10³ L/mol, 2.47, respectively. Thermodynamic parameters including enthalpy change (ΔH) and entropy change (ΔS) were calculated, the results suggesting that hydrophobic force played a very important role in SD‐Na binding to HSA, which was in good agreement with the molecular modeling study. Moreover, the effect of SD‐Na on the conformation of HSA was analyzed using three‐dimensional fluorescence spectra. Copyright © 2013 John Wiley & Sons, Ltd.