Hydrophilic carotenoids: surface properties and aggregation behavior of a highly unsaturated carotenoid lysophospholipid

The water dispersibility of a hydrophobic carotenoid has been greatly enhanced by using it as the acyl part in the synthesis of a highly unsaturated lysophospholipid. Dynamic light scattering has revealed the formation of stable aggregates with an average hydrodynamic radius of a few nanometers, and absorption spectra show that the aggregates can withstand the addition of ethanol or acetonitrile until the volume fraction of water falls below 70 and 62%, respectively. The properties of the carotenoid phospholipids have been characterized by determining surface tension, critical micelle concentration, surface concentration, molecular area, free energy of adsorption and micellation, adsorption-micellar energy relationship, and equilibrium constants.     

Discovery of human inversion polymorphisms by comparative analysis of human and chimpanzee DNA sequence assemblies

With a draft genome-sequence assembly for the chimpanzee available, it is now possible to perform genome-wide analyses to identify, at a submicroscopic level, structural rearrangements that have occurred between chimpanzees and humans. The goal of this study was to investigate chromosomal regions that are inverted between the chimpanzee and human genomes. Using the net alignments for the builds of the human and chimpanzee genome assemblies, we identified a total of 1,576 putative regions of inverted orientation, covering more than 154 mega-bases of DNA. The DNA segments are distributed throughout the genome and range from 23 base pairs to 62 mega-bases in length. For the 66 inversions more than 25 kilobases (kb) in length, 75% were flanked on one or both sides by (often unrelated) segmental duplications. Using PCR and fluorescence in situ hybridization we experimentally validated 23 of 27 (85%) semi-randomly chosen regions; the largest novel inversion confirmed was 4.3 mega-bases at human Chromosome 7p14. Gorilla was used as an out-group to assign ancestral status to the variants. All experimentally validated inversion regions were then assayed against a panel of human samples and three of the 23 (13%) regions were found to be polymorphic in the human genome. These polymorphic inversions include 730 kb (at 7p22), 13 kb (at 7q11), and 1 kb (at 16q24) fragments with a 5%, 30%, and 48% minor allele frequency, respectively. Our results suggest that inversions are an important source of variation in primate genome evolution. The finding of at least three novel inversion polymorphisms in humans indicates this type of structural variation may be a more common feature of our genome than previously realized.     

Characterization of growth hormone binding sites in granulosa and theca layers at different stages of follicular maturation and ovulatory cycle in the domestic hen

The currently available evidence points to a possible influence of growth hormone (GH) on avian folliculogenesis, which can be mediated by both hepatic- and ovarian-derived IGF-I. Therefore, the purpose of the present study was to reveal GH-binding sites in granulosa and theca layers of preovulatory follicles and to determine the binding characteristics depending on the degree of follicular maturation and the stage of the ovulatory cycle in the hen. Hens were killed 2 h (stage I), 9 h (stage II), 16 h (stage III), and 23 h (stage IV) after oviposition, and the five largest yellow follicles (from F1 to F5) were removed. GH-binding sites in granulosa and theca layers from F1 to F5 follicles were characterized using a radioreceptor assay. Equilibrium dissociation constants (K(d)) and binding capacities (B(max)) were determined by Scatchard analysis of saturation curves, which revealed a single class of high-affinity GH-binding sites in both theca tissue and granulosa cells. In F1, F2, and F5 follicles, B(max) and K(d) for GH-binding sites in the granulosa layer changed during the ovulatory cycle, decreasing between stages I and III, to increase again at stage IV, with alterations in K(d) being less profound. No significant differences in binding capacities and affinities of GH-binding sites in the theca layer were found between various stages of the cycle. Furthermore, the concentration of GH-binding sites in the granulosa layer rose, whereas that in the theca layer fell with follicular enlargement. These findings indicate the presence of high-affinity GH-binding sites in both granulosa and theca layers of hen preovulatory follicles. Data also demonstrate that GH-binding sites in these tissues are regulated in a tissue-specific manner. Furthermore, the regulation of binding capacity of GH binding in granulosa cells by hormonal factors associated with ovulatory cycle is apparently not dependent on the state of follicular maturation.     

Comparative study of integrating cavity absorption meters

Integrating spheres (IS) are widely used for recording spectra of scattering samples by placing the specimen inside or outside the sphere. An unusual application of integrating spheres has been also demonstrated earlier where the liquid sample completely filled the spherical cavity; such a device is often called an integrating cavity absorption meter (ICAM). In the present work, integrating cavities with different coatings are compared. The spheres were made of glass, covered by metal or white paint, and their surfaces were prepared for diffuse or specular reflectance. The spheres were evaluated by recording kinetic traces following a short light pulse with the aid of time-correlated single photon counting (TCSPC), and by recording steady-state spectra through single-photon counting (SPC) detection. The relative efficiencies of the spheres were determined by comparing the steady-state spectra. Possible reasons for differences in the performance are discussed.     

Heteroaromatic-aminomethyl quinolones: potent and selective iNOS inhibitors

The overproduction of nitric oxide during the biological response to inflammation by the nitric oxide synthase (NOS) enzymes have been implicated in the pathology of many diseases. By removal of the amide core from uHTS-derived quinolone 4, a new series highly potent heteroaromatic-aminomethyl quinolone iNOS inhibitors 8 were identified. SAR studies led to identification of piperazine 22 and pyrimidine 32, both of which reduced plasma nitrates following oral dosing in a mouse lipopolysaccharide challenge assay.     

DNA interaction studies of a copper (II) complex containing an antiviral drug, valacyclovir: the effect of metal center on the mode of binding

The water-soluble complex, [Cu(Val)(2)(NO(3))(2)]; in which Val = valacyclovir, an antiviral drug, has been synthesized and characterized by elemental analysis, furier transfer-infrared, hydrogen nuclear magnetic resonance (H NMR), and UV-Vis techniques. The binding of this Cu (II) complex to calf thymus DNA was investigated using fluorimetry, spectrophotometry, circular dichroism, and viscosimetry. In fluorimetric studies, the enthalpy and entropy of the reaction between the complex and calf-thymus DNA (CT-DNA) showed that the reaction is endothermic (ΔH = 208.22 kJ mol(-1); ΔS = 851.35 J mol(-1)K(-1)). The complex showed the absorption hyperchromism in its ultra violet-visible (UV-Vis) spectrum with DNA. The calculated binding constant, K(b), obtained from UV-Vis absorption studies was 2 × 10(5) M(-1). Moreover, the complex induced detectable changes in the circular dichroism spectrum of CT-DNA, as well as changes in its viscosity. The results suggest that this copper (II) complex interacts with CT-DNA via a groove-binding mode.     

Interaction of calf thymus DNA with the antiviral drug Lamivudine

One approach to accelerate the availability of new cancer drugs is to test drugs approved for other conditions as anticancer agents. In recent years, some researchers have shown that antiviral drugs, such as ritonavir, saquinavir, and nelfinavir, inhibit the growth of over 60 cancer cell lines derived from nine different tumor types. This article studied the anticancer potential of an antiviral drug, lamivudine (LA). The interaction of LA and calf thymus DNA (CT-DNA) was studied using emission, absorption, circular dichroism (CD), and viscosity techniques. The binding constants evaluated from fluorescence data at different temperatures revealed that fluorescence enhancement is a static process that involves complex-DNA formation in the ground state. Further, the enthalpy and entropy of the reaction between the drug and CT-DNA showed ΔH<0 (-126.38±0.61 kJ mol(-1)) and ΔS<0 (-352.17±2.1?J mol(-1) K(-1)); therefore, van der Waals interactions or hydrogen bonds are the main forces in the binding of LA to CT-DNA. The values of K(f) clearly underscore the high affinity of LA to DNA. In addition, detectable changes in the CD spectrum of CT-DNA in the presence of LA indicated conformational changes. All these results showed that groove binding is the binding mode of this drug and CT-DNA.     

Use of allele-specific sequencing primers is an efficient alternative to PCR subcloning of low-copy nuclear genes

Direct Sanger sequencing of polymerase chain reaction (PCR)-amplified nuclear genes leads to polymorphic sequences when allelic variation is present. To overcome this problem, most researchers subclone the PCR products to separate alleles. An alternative is to directly sequence the separate alleles using allele-specific primers. We tested two methods to enhance the specificity of allele-specific primers for use in direct sequencing: using short primers and amplification refractory mutation system (ARMS) technique. By shortening the allele-specific primer to 15-13 nucleotides, the single mismatch in the ultimate base of the primer is enough to hinder the amplification of the nontarget allele in direct sequencing and recover only the targeted allele at high accuracy. The deliberate addition of a second mismatch, as implemented in the ARMS technique, was less successful and seems better suited for allele-specific amplification in regular PCR rather than in direct sequencing.     

Light absorption by anthocyanins in juvenile, stressed, and senescing leaves

The optical properties of leaves from five species, Norway maple (Acer platanoides L.), cotoneaster (Cotoneaster alaunica Golite), hazel (Corylus avellana L.), Siberian dogwood (Cornus alba L.), and Virginia creeper (Parthenocissus quinquefolia (L.) Planch.), differing in pigment composition and at different stages of ontogenesis, were studied. Anthocyanin absorption maxima in vivo, as estimated with spectrophotometry of intact anthocyanic versus acyanic leaves and microspectrophotometry of vacuoles in the leaf cross-sections, were found between 537 nm and 542 nm, showing a red shift of 5-20 nm compared with the corresponding maxima in acidic water-methanol extracts. In non-senescent leaves, strong anthocyanin absorption was found between 500 nm and 600 nm (with a 70-80 nm apparent bandwidth). By and large, absorption by anthocyanin in leaves followed a modified form of the Lambert-Beer law, showing a linear trend up to a content of nearly 50 nmol cm(-2), and permitting thereby a non-invasive determination of anthocyanin content. The apparent specific absorption coefficients of anthocyanins at 550 nm showed no substantial dependence on the species. Anthocyanin contribution to total light absorption at 550 nm was followed in maple leaves in the course of autumn senescence. Photoprotection by vacuolar anthocyanins is discussed with special regard to their distribution within a leaf; radiation screening by anthocyanins predominantly localized in the epidermal cells in A. platanoides and C. avellana leaves was also evaluated.     

NK cell protease granzyme M targets alpha-tubulin and disorganizes the microtubule network

Serine protease granzyme M (GrM) is highly expressed in the cytolytic granules of NK cells, which eliminate virus-infected cells and tumor cells. The molecular mechanisms by which GrM induces cell death, however, remain poorly understood. In this study we used a proteomic approach to scan the native proteome of human tumor cells for intracellular substrates of GrM. Among other findings, this approach revealed several components of the cytoskeleton. GrM directly and efficiently cleaved the actin-plasma membrane linker ezrin and the microtubule component alpha-tubulin by using purified proteins, tumor cell lysates, and tumor cells undergoing cell death induced by perforin and GrM. These cleavage events occurred independently of caspases or other cysteine proteases. Kinetically, alpha-tubulin was more efficiently cleaved by GrM as compared with ezrin. Direct alpha-tubulin proteolysis by GrM is complex and occurs at multiple cleavage sites, one of them being Leu at position 269. GrM disturbed tubulin polymerization dynamics in vitro and induced microtubule network disorganization in tumor cells in vivo. We conclude that GrM targets major components of the cytoskeleton that likely contribute to NK cell-induced cell death.