Strand Structure Research Articles

Eliminating the secondary structure of targeting strands for enhancement of DNA probe based low-abundance point mutation detection

Nucleic acid probes are very useful tools in biological and medical science. However, the essential sensing mechanism of nucleic acid probes was prone to the interference of surrounding sequences. Especially when the target sequences formed secondary structures such as hairpin or quadruplex, the nucleic acid probes were hindered from hybridizing with target strands, greatly disabled the function of probes. Herein, we have established an Open strand based strategy for eliminating the influence of secondary structures on the performance of nucleic acid probes. The strategy was general toward different lengths, secondary structures and sequences of the targeting strand, and we found that the improvement was higher when the secondary structure of the targeting strand was more complicated. Experiments on synthetic single stranded DNA and real clinical genomic DNA samples were conducted for low abundance mutation detection, and the limit of detection for TERT-C228T and BRCA2 rs80359065 mutations could be 0.02% and 0.05% respectively, demonstrating the clinical practicability of our proposed strategy in low abundance mutation detection.

Surface Glycoproteomic Analysis Reveals That Both Unique and Differential Expression of Surface Glycoproteins Determine the Cell Type.

Proteins on the cell surface are frequently glycosylated, and they are essential for cells. Surface glycoproteins regulate nearly every extracellular event, but compared with global analysis of proteins, comprehensive and site-specific analysis of surface glycoproteins is much more challenging and dramatically understudied. Here, combining metabolic labeling, click-chemistry and enzymatic reactions, and mass spectrometry-based proteomics, we globally characterized surface glycoproteins from eight popular types of human cells. This integrative and effective method allowed for the identification of 2172 N-glycosylation sites and 1047 surface glycoproteins. The distribution and occurrence of N-glycosylation sites were systematically investigated, and protein secondary structures were found to have a dramatic influence on glycosylation sites. As expected, most sites are located on disordered regions. For the sites with the motif N-!P-C, about one-third of them are located on helix structures, while those with the motif N-!P-S/T prefer strand structures. There is almost no correlation between the number of glycosylation sites and protein length, but the number of sites corresponds well with the frequencies of the motif. Quantification results reveal that besides cell-specific glycoproteins, the uniqueness of each cell type further arises from differential expression of surface glycoproteins. The current research indicates that multiple surface glycoproteins including their abundances need to be considered for cell classification rather than a single cluster of differentiation (CD) protein normally used in conventional methods. These results provide valuable information to the glycoscience and biomedical communities and aid in the discovery of surface glycoproteins as disease biomarkers and drug targets.

A Fluorescence Light-Up Silver Nanocluster Beacon Modulated by Metal Ions and Its Application in Telomerase-Activity Detection.

DNA-templated silver nanoclusters (DNA-AgNCs) have been extensively studied in recent years. The enhancement of fluorescence emission from DNA-AgNCs is still being explored. Herein, a study on the fluorescence enhancement of DNA-AgNCs induced by metal ions is reported. The enhancement is greatly dependent on the primary sequence and secondary structure of the DNA strands. Thus, a label-free AgNCs-based molecular beacon (MB) was explored for the detection of telomerase activity. Nonfluorescent MB-AgNCs in phosphate buffer emit dramatic red fluorescence when Mg2+ is introduced, whereas Mg2+ has a limited effect on the weak fluorescence of DNA-AgNCs when the hairpin structure of MB is opened. Telomerase primer can be elongated by telomerase, which results in unfolding of MB in a strand-displacement reaction. On the basis of the different brightnesses of AgNCs produced by the two DNA templates, telomerase activity can be detected. The MB-AgNCs sensing platform provides a simple and low-cost method to detect telomerase activity and shows great potential in the construction of cost-effective probes for biomolecular detection.

Lanthanide Coordination Polymers and MOFs based on the Dicyanodihydridoborate Anion.

New lanthanide cyanoborates were synthesized from anhydrous lanthanide chlorides and the acid H[BH2 (CN)2 ] in either acetonitrile or pyridine. Reactions in acetonitrile lead to three-dimensional, anionic metal-organic frameworks (MOFs) 3 ∞ [Ln2 {BH2 (CN)2 }9 ]⋅[Ln(CH3 CN)9 ] (Ln=Ce, Eu, Tb) which incorporate complex cations [Ln(CH3 CN)9 ]3+ in the pores of the framework for charge compensation. In contrast, the reactions in pyridine result in the formation of one-dimensional coordination polymers 1 ∞ [H(py)2 ][LnCl2 {BH2 (CN)2 }2 (py)2 ]⋅0.5 py (Ln=Ce, Pr, py=pyridine) with [H(py)2 ]+ as counter ions for the anionic strand structure. The products show intense photoluminescence, for Ce3+ based on 5d-4f transitions in the blue spectral region, whereas the Eu3+ and Tb3+ compounds exhibit characteristic photoluminescence based on 4f-4f transitions of the respective lanthanide ions. The observed photoluminescence is mainly attributed to a direct excitation of the lanthanide ions and sensitization of the lanthanide ions by the [BH2 (CN)2 ]- anions. These results mark the utilized borate anions as versatile building block for new coordination compounds.

Quantitative descriptions of extended volume cardiac tissue architecture from multiple large hearts.

The arrangement of cardiac cells into strand and sheet-like structures within the heart wall, confers important electrical properties onto heart tissue. Unraveling cardiomyocyte architecture in both healthy and diseased hearts is fundamental to understanding the mechanisms generating normal rhythm and arrhythmia. We analyzed five extended volume serial image stacks of normal pig left ventricular tissue. Analysis included: (1) reconstruction of original tissue volume and shape with non-linear correction maps; (2) segmentation and higher-order descriptions, areas and orientations of laminar structures through the heart wall; (3)computation of fiber directions; (4) computation of tissue connectivity using a shell filter. These measures contributed to a deeper and more objective understanding of cardiac tissue structures and their spatial variation than previously possible.

Amino acid content of beta strands and alpha helices depends on their flanking secondary structure elements

The influence of flanking structures (alpha helices and beta strands in the primary sequence) on amino acid content of the elements of secondary structure has been analyzed in seven sets of nonhomologous proteins. Elevated usage of beta structural amino acid residues and pentapeptides in beta strands between two alpha helices can be explained by the stabilization of secondary structure of those beta strands by natural selection. High usage of alpha helical amino acids and pentapeptides in beta strands situated between two other beta strands is an evidence of the relaxation of natural selection: “passive” beta strands in these fragments of polypeptide chains are frequently formed due to the influence of flanking “active” beta strands. Alpha helices situated between alpha helix and beta strand are enriched by alpha helical pentapeptides and have lower usage of beta structural pentapeptides than those situated between beta strand and alpha helix, their N-termini are more frequently protected from helix to beta transitions by Glu residues. Alpha helices between two beta strands are stabilized by Ala residues and pentapeptides specific to alpha helices. Dipeptide content of the most stable alpha helices and beta strands was used for the creation of the PentaFOLD 2.0 algorithm (http://chemres.bsmu.by) that finds stable fragments of these elements of secondary structure in PDB files.

Marriage Reform and the Constitution of the United Reformed Church

Recent reforms to English and Scots marriage law faced the United Reformed Church (URC) with two challenges. Its hybrid structure of church government, entwining Congregational and Presbyterian strands, complicated application of the statutory criterion ‘persons recognised by [the membership] as competent for the purpose of giving consent’. Precedent from earlier decisions on human sexuality explains the ultimate identification of the local church meeting as the competent council of the URC in England, and why the ‘enabling resolution’ passed regarding civil partnership formation was not repeated. The very different focus of Scots marriage law posed different questions, less focused on buildings or the churches using them and allowing willing celebrants to be nominated by the synod, as for opposite-sex marriage.Advisers differed on whether the denomination possessed any binding doctrine of marriage which would obstruct implementation of the amended law. The General Assembly decision on polity and how it was reached suggest an implicit ruling in the negative. This article defends that outcome, considering the doctrinal foundation of the URC in the light of concessions made at the formative union. Marriage appears as a topic on which no denominational doctrine exists, letting all councils reach theological conclusions necessary to practical decisions within their remit.

Proflavine an acridine DNA intercalating agent and strong antimicrobial possessing potential properties of carcinogen

Proflavine finds a wide array of applications in clinical, therapeutic, industrial and cutting edge research. This study investigates properties of proflavine and brings out its current status based on overall merits and demerits. Review was carried out starting from late 1800s to 2017 about all aspects of proflavine. We have accessed digital libraries in University of Cambridge, Oxford University, Harvard University and Massachusetts Institute of Technology through their institutional repository software. Popular open source solutions like DSpace, EPrints, Digital Commons, Fedora Commons, Islandora and Hydra were included. Proflavine finds many applications including as an anti microbial agent and often used as a topical agent. This compound denatures bacterial DNA leading to lysis of bacteria. Due to its intercalating property it affects host DNA, which has potential chances to induce skin cancer and other malignancies. Reactive oxygen species (ROS) released by proflavine play a crucial role in denaturing host DNA. Proflavine can penetrate beyond epidermal and dermal structures and accumulate in cell nuclei. In human cell culture proflavine is known to be taken up by many kinds of cells and is concentrated in the nuclei. Our studies revealed that despite its oncogenic potential, proflavine currently finds its applications in therapeutic, diagnostic and in research not alone in developing countries but also in developed countries. Proflavine exhibited wide potent activity against various groups of microorganisms. After exposure in human skin proflavine alters the structure of epidermal DNA strands leading to mutation. Many industrial workers, researchers, health care professionals are exposed to proflavine in enormous amounts daily through oral, respiratory and cutaneous routes. We conclude that even though proflavine has strong antimicrobial and other uses due to its carcinogenic nature, we recommend that it is time to rethink the use of this compound and to search for good alternative replacement.

Interaction of sucralose with whey protein: Experimental and molecular modeling studies

The objective of this research was to study the interactions of sucralose with whey protein isolate (WPI) by using the three-dimensional fluorescence spectroscopy, circular dichroism spectroscopy and molecular modeling. The results showed that the peptide strands structure of WPI had been changed by sucralose. Sucralose binding induced the secondary structural changes and increased content of aperiodic structure of WPI. Sucralose decreased the thermal stability of WPI and acted as a structure destabilizer during the thermal unfolding process of protein. In addition, the existence of sucralose decreased the reversibility of the unfolding of WPI. Nonetheless, sucralose–WPI complex was less stable than protein alone. The molecular modeling result showed that van der Waals and hydrogen bonding interactions contribute to the complexation free binding energy. There are more than one possible binding sites of WPI with sucralose by surface binding mode.

Habitat structure and zonation patterns of northwestern Mediterranean shoreline strands

We studied the habitat structure (macrofaunal assemblages and bottom types) and zonation patterns of 29 unvegetated shoreline strands along the 900-km coast of Catalonia (NW Mediterranean Sea). Organisms were sampled with grabs, pitfall traps, sticky traps, clam nets and spades to ensure capture of the different proportions of macrofaunal assemblages from the supra-, medio- and infralittoral levels. We collected 211 taxa: 194 animals and 17 algae. The most abundant and dominant organisms collected with van Veen grabs were Nematoda, Oligochaeta and Collembola at the supralittoral level; the polychaetes Saccocirrus spp. and Pisione remota, the amphipod Corophium orientale, Nematoda, and Turbellaria at the mediolittoral level; and Nematoda at the upper infralittoral level. SIMPER analysis revealed great dissimilarity between the organisms inhabiting the supralittoral and the other littoral levels. Regarding the epifauna, the sticky traps used at the supralittoral level mainly collected Collembola, which were nearly absent in pitfall traps. The qualitative study performed with a clam net and a small spade revealed that Nematoda, Saccocirrus spp., Turbellaria, Nemertea and the polychaete P. remota were the most abundant animals at both the medio- and the infralittoral levels and no differences were found between these levels. Different qualitative sampling methodologies showed that in fine sediments the bivalves Donax trunculus and D. semistriatus determined more than 97% of dissimilarity from coarse-sand sites. Richness increased in protected sandy and cobble shores. Littoral level and bottom-type features were only to a certain extent valid indicators of specific biotic components for a specific habitat.

A mechanistic study of helicases with magnetic traps.

Helicases are a broad family of enzymes that separate nucleic acid double strand structures (DNA/DNA, DNA/RNA, or RNA/RNA) and thus are essential to DNA replication and the maintenance of nucleic acid integrity. We review the picture that has emerged from single molecule studies of the mechanisms of DNA and RNA helicases and their interactions with other proteins. Many features have been uncovered by these studies that were obscured by bulk studies, such as DNA strands switching, mechanical (rather than biochemical) coupling between helicases and polymerases, helicase-induced re-hybridization and stalled fork rescue.

Discrimination of Polynucleotide Transport through a Highly Hydrophobic Uncharged Nanopore

The importance of DNA structures in medicine has motivated researchers to develop sequencing methods with low costs and high speeds. The potential use of nanofluidics for the analysis of the translocation of polynucleotides through a nanopore is a very interesting solution. Herein, we demonstrate both by experiments and by explicit solvent molecular dynamics simulations that the translocation of polyadenylic acid and polycytidylic acid can significantly affect the ionic flux behavior. In particular, the nanochannel enhances the velocity difference between polynucleotides during the translocation process. Our results revealed the important role of the strand structures during their translocation, as already observed in biochannels. The folding/unfolding process leads to stronger interactions with the nanopore wall that decrease the velocity of the strands. This behavior could lead to the detection of different polynucleotides in a mixture.

Mn(II), Zn(II) and Cd(II) Complexes Based on Oxadiazole Backbone Containing Carboxyl Ligand: Synthesis, Crystal Structure, and Photoluminescent Study.

Three coordination polymers, [Cd(L)2(H2O)2]n (1), [Zn(L)2(H2O)2]n (2) and [Mn(L)2]n (3) were prepared by reacting 5-(3-pyridyl)-1,3,4-oxadiazole-2-thioacetic acid (HL) with corresponding metal acetate in DMF/CH3CN medium under solvothermal condition. The isolated complexes were characterized by elemental analysis and infrared spectroscopy. The X-ray crystallographic analysis revealed double strand structure of 1 and 2, and 3D framework of 3. The different structures of these complexes indicate that the configuration of the ligand and the reaction condition play a key role in self-assemble of complexes 1-3. Furthermore, photoluminescent properties of 1 and 2 were also studied in the solid state.

How base pairs stack up

Biophysics The interactions between nucleic acid bases include not only the hydrogen bonding between bases but also the π-orbital interactions between base pairs. To probe these interactions, Kilchherr et al. constructed an ensemble of DNA origami structures of blunt-end DNA strands. The forces in

Strand withdrawal rate stabilization: via the electric drive of the secondary cooling zone of a continuous casting machine

Statement of the problem: stabilization of the rate of strand withdrawal out of the mould and the secondary cooling zone after change operations of a tundish or a submerged nozzle is one of the attempts to increase the productivity of a continuous casting machine and the quality of a casting strand. The aim of this work is to study the causes of oscillation occurrence in the rate of strand withdrawal and the development of the electric drive control system of the secondary cooling zone providing the increased machine productivity via stabilization of the rate at preserving the required quality of the internal structure of cast strands. Experimental studies were based on the analysis of the changes in load currents of withdrawal roll motors, the total current of the electric drive of the secondary cooling zone of continuous casting machines, macro template laboratory data on the main internal defects detected in strands with the use of methods of statistical data processing. The result: the method proposed for stabilization of the rate of strand withdrawal by means of compensating the harmonic component of the total moment of resistance of strand withdrawal by the reduced amplitude of oscillations of the strand withdrawal rate more than three times allows improving the quality of the macrostructure of cast strands with the increased strand withdrawal rate by 5 %. The results of mathematical modeling and experimental studies can be used in the design of automated electric drives of the secondary cooling zone of continuous casting machines of curved type.

A role for loop G in the β1 strand in GABAA receptor activation.

The role of the β1 strand in GABAA receptor function is unclear. It lies anti-parallel to the β2 strand, which is known to participate in receptor activation. Molecular dynamics simulation revealed solvent accessible residues within the β1 strand of the GABAA β3 homopentamer that might be amenable to analysis using the substituted Cys accessibility method. Cys substitutions from Asp43 to Thr47 in the GABAA α1 subunit showed that D43C and T47C reduced the apparent potency of GABA. F45C caused a biphasic GABA concentration-response relationship and increased spontaneous gating. Cys43 and Cys47 were accessible to 2-aminoethyl methanethiosulphonate (MTSEA) modification, whereas Cys45 was not. Both GABA and the allosteric agonist propofol reduced MTSEA modification of Cys43 and Cys47. By contrast, modification of Cys64 in the β2 strand loop D was impeded by GABA but unaffected by propofol. These data reveal movement of β1 strand loop G residues during agonist activation of the GABAA receptor. The GABAA receptor α subunit β1 strand runs anti-parallel to the β2 strand, which contains loop D, known to participate in receptor activation and agonist binding. However, a role for the β1 strand has yet to be established. We used molecular dynamics simulation to quantify the solvent accessible surface area (SASA) of β1 strand residues in the GABAA β3 homopentamer structure. Residues in the complementary interface equivalent to those between Asp43 and Thr47 in the α1 subunit have an alternating pattern of high and low SASA consistent with a β strand structure. We investigated the functional role of these β1 strand residues in the α1 subunit by individually replacing them with Cys residues. D43C and T47C substitutions reduced the apparent potency of GABA at α1β2γ2 receptors by 50-fold and eight-fold, respectively, whereas the F45C substitution caused a biphasic GABA concentration-response relationship and increased spontaneous gating. Receptors with D43C or T47C substitutions were sensitive to 2-aminoethyl methanethiosulphonate (MTSEA) modification. However, GABA-evoked currents mediated by α1(F45C)β2γ2 receptors were unaffected by MTSEA, suggesting that this residue is inaccessible. Both GABA and the allosteric agonist propofol reduced MTSEA modification of α1(D43C)β2γ2 and α1(T47C)β2γ2 receptors, indicating movement of the β1 strand even during allosteric activation. This is in contrast to α1(F64C)β2γ2 receptors, where only GABA, but not propofol, reduced MTSEA modification. These findings provide the first functional evidence for movement of the β1 strand during gating of the receptor and identify residues that are critical for maintaining GABAA receptor function.

Unique infection structures produced by Pseudocercosporella capsellae on oilseed crops Brassica carinata, B. juncea and B. napus in Western Australia

White leaf spot disease (Pseudocercosporella capsellae) is widespread across oilseed, vegetable and forage brassicas. Light (LM) and scanning electron (SEM) microscope studies were undertaken to investigate host–pathogen interactions on cotyledons of resistant and susceptible Brassica carinata, B. juncea and B. napus. Under LM, unique brown structures were present, particularly on susceptible genotypes, in two morphologically distinct forms: first, as thread‐like structures within cortical tissue by 24 h post‐inoculation (hpi) and secondly, as brown ropy strand structures either within cortical tissues (internal ropy strands), or extruded out through stomatal pores (ropy strand extrusions). Under LM, these brown structures were most prevalent in highly susceptible B. juncea ‘Vardan’ that had both a high incidence within cortical tissue (70%) and of ropy strand extrusions (73%), as did susceptible B. napus ‘Trilogy’ within cortical tissue (60%). Under SEM, both these genotypes showed thread‐like structures smaller than hyphae forming highly branched networks and ropy strand‐like structures. While there were fewer brown structures in susceptible B. carinata UWA #012 (35%), fine, thread‐like structures forming networks were again prominent (SEM). In contrast, for resistant genotypes, brown structures (LM) were of very low frequency or absent; only 5% in resistant B. juncea ‘Dune’ and none in resistant B. napus ‘Hyola 42’ or highly resistant B. carinata ATC94129P. Under SEM, fine, thread‐like structures were present in the resistant B. juncea ‘Dune’ and B. napus ‘Hyola 42’. Liquid chromatographic analyses of brown structures revealed that both internal ropy strands within cortical tissues and ropy strand extrusions contained the mycotoxin cercosporin.

Enhanced DNA imaging using super-resolution microscopy and simultaneous single-molecule orientation measurements.

Single-molecule orientation measurements provide unparalleled insight into a multitude of biological and polymeric systems. We report a simple, high-throughput technique for measuring the azimuthal orientation and rotational dynamics of single fluorescent molecules, which is compatible with localization microscopy. Our method involves modulating the polarization of an excitation laser, and analyzing the corresponding intensities emitted by single dye molecules and their modulation amplitudes. To demonstrate our approach, we use intercalating and groove-binding dyes to obtain super-resolved images of stretched DNA strands through binding-induced turn-on of fluorescence. By combining our image data with thousands of dye molecule orientation measurements, we develop a means of probing the structure of individual DNA strands, while also characterizing dye-DNA interactions. This approach may hold promise as a method for monitoring DNA conformation changes resulting from DNA-binding proteins.

Structural characterization of complexes prepared with glycerol monoestearate and maize starches with different amylose contents

Morphology and different structural features of V-amylose complexes prepared with different concentrations (1%, 2%, 3%) of glycerol monostearate (GMS) and normal maize (NMS), waxy maize (WMS), and high amylose maize (HAMS) starches were evaluated using X-ray diffraction, differential scanning calorimetry, scanning electronic microscopy (SEM), atomic force microscopy (AFM), and transmission electronic microscopy (TEM). There was inclusion complex formation between all starches and GMS regardless of emulsifier concentration, with exception of WMS-2%GMS and WMS-3%GMS samples. All of the inclusion complexes displayed a V-type crystalline pattern and endothermic dissociation peaks between 115 and 120°C. They also displayed faceted crystalline structures with a tendency of the crystals to aggregate and form agglomerates of various sizes. TEM images of the complexes showed an aggregated strand structure interwoven with the GMS. Emulsifier and amylose quantities directly influenced complex formation. At high GMS concentrations, there was higher tendency of emulsifier to self-associate rather than form complexes with amylose.

Synthesis, structure and properties of cholesterol-based A(LS)2- and A(LS)3-type gelators without hydrogen bond linkers

Dicholesteryl isophthalate (1m), dicholesteryl terephthalate (1p) and tricholesteryl trimesate (2) were synthesized as new cholesterol-based A(LS)n-type gelators (n=2 and 3) without hydrogen bond linkers. Their molecular structural details were investigated experimentally by 1H and 13C NMR, UV–vis and IR spectroscopies and theoretically by DFT calculations. Compounds 1p and 2 have relatively high degrees of symmetry and exhibit efficient gelation abilities, whereas compound 1m could not be gelated in any common organic solvents. The self-assembly behavior of the compound was largely affected by the type of solvent and the number and substitution positions of the cholesteryl groups, as revealed by scanning electron microscopy (SEM) measurements. Notably, the xerogel of 2 using benzene displayed an aggregated microfibrous strand structure that included helical fibers.