Single Helical Conformation Research Articles

Single-helical formyl β-glucan effectively deliver CpG DNA with poly(dA) to macrophages for enhanced vaccine effects

Single helical β-glucan is a one-dimensional host that can form a hybrid helix with DNAs/RNAs as delivery systems. However, unmodified β-glucan has a gelling tendency and a single helical conformation is challenging to obtain. Therefore, in this study, we developed a β-glucan formyl derivative with stable single helical conformation and no gelling tendency. Circular dichroism studies found that the formyl-β-glucan could form a hybrid helix with DNA CpG-poly(dA). The hybrid helix delivery system showed improved activation on antigen-presenting cells, thereby upregulating the mRNA and protein levels of inflammatory factors, and had an immune-enhancing effect on ovalbumin (OVA) immunized mice. These results indicate that formyl-β-glucan can be developed as a non-cationic supramolecular DNA delivery platform with low toxicity and high efficiency.

Tunable Circularly Polarized Luminescence from Single Crystal and Powder of the Simplest Tetraphenylethylene Helicate.

Tetraphenylethylene and its derivatives are a class of aggregation-induced emission (AIE) compounds that are most extensively and successfully studied. It has been found that the simplest TPE is easy to crystallize into homochiral M crystals or P crystals. However, no research on circularly polarized luminescence (CPL) of TPE solid is documented. In this paper, we report that TPE can grow into big and nonefflorescent single crystals in single helical conformation and has large birefringence that is comparative with commercially available products. The TPE single crystals can emit strong CPL with a very high glum value up to 0.53. Moreover, the sense and magnitude of CPL signals can be willfully tuned by simple rotation of the single crystal due to anisotropy of the crystals. This simple and promising CPL photonic material integrates emission, chirality, and birefringence together in one single crystal without needing an additional chiral dopant or conjugate polymer that can produce linearly polarized light. After being ground into fine powder and pressed as KBr pellets, the obtained nanocrystals of TPE also emit strong CPL light. Exceptionally, by mixing other achiral luminescent dyes together with TPE powder in KBr pellets, induced CPL signals were obtained, which could give full-color CPL emission. Although there were no interactions between TPE and the dyes in the pellets, induced CPL signals were realized through radiative energy transfer, providing a very simple method for the tuning of CPL emission.

Internalization of Foldamer-Based DNA Mimics through a Site-Specific Antibody Conjugate to Target HER2-Positive Cancer Cells.

Inhibition of protein–DNA interactions represents an attractive strategy to modulate essential cellular functions. We reported the synthesis of unique oligoamide-based foldamers that adopt single helical conformations and mimic the negatively charged phosphate moieties of B-DNA. These mimics alter the activity of DNA interacting enzymes used as targets for cancer treatment, such as DNA topoisomerase I, and they are cytotoxic only in the presence of a transfection agent. The aim of our study was to improve internalization and selective delivery of these highly charged molecules to cancer cells. For this purpose, we synthesized an antibody-drug conjugate (ADC) using a DNA mimic as a payload to specifically target cancer cells overexpressing HER2. We report the bioconjugation of a 16-mer DNA mimic with trastuzumab and its functional validation in breast and ovarian cancer cells expressing various levels of HER2. Binding of the ADC to HER2 increased with the expression of the receptor. The ADC was internalized into cells and was more efficient than trastuzumab at inhibiting their growth in vitro. These results provide proof of concept that it is possible to site-specifically graft high molecular weight payloads such as DNA mimics onto monoclonal antibodies to improve their selective internalization and delivery in cancer cells.

Preparation and characterization of curdlan with unique single-helical conformation and its assembly with Congo Red

Elucidating the structure-activity relationship of curdlan is hampered by a lack of characterization with unique specific conformations (i.e., single- or triple-helix). In this study, single-helical curdlan is generated in dilute NaOH solutions at 35−50 °C, and characterized with NMR, SAXS, and GPC. The conformational transition from coil to single-helix and the intramolecular hydrogen bond interaction are explored using NMR. It is found that the two aforementioned types of curdlan interact with Congo Red in very different ways. Single-helical curdlan can encapsulate Congo Red to form a stable, supramolecular dye assembly, which is demonstrated by the shortest distance between the H3 of curdlan and the phenyl groups of Congo Red, and also the same self-diffusion coefficients of Congo Red and curdlan. In contrast, random-coil curdlan interacts weakly with Congo Red and cannot enwrap it. This study offers insight into the specific structure-activity relationship of beta-(1,3)-glucans.

Single helically folded aromatic oligoamides that mimic the charge surface of double-stranded B-DNA.

Numerous essential biomolecular processes require the recognition of DNA surface features by proteins. Molecules mimicking these features could potentially act as decoys and interfere with pharmacologically or therapeutically relevant protein-DNA interactions. Although naturally occurring DNA-mimicking proteins have been described, synthetic tunable molecules that mimic the charge surface of double-stranded DNA are not known. Here, we report the design, synthesis and structural characterization of aromatic oligoamides that fold into single helical conformations and display a double helical array of negatively charged residues in positions that match the phosphate moieties in B-DNA. These molecules were able to inhibit several enzymes possessing non-sequence-selective DNA-binding properties, including topoisomerase 1 and HIV-1 integrase, presumably through specific foldamer-protein interactions, whereas sequence-selective enzymes were not inhibited. Such modular and synthetically accessible DNA mimics provide a versatile platform to design novel inhibitors of protein-DNA interactions.

Structural Advantage of Sugar Beet α-Glucosidase to Stabilize the Michaelis Complex with Long-chain Substrate

The α-glucosidase from sugar beet (SBG) is an exo-type glycosidase. The enzyme has a pocket-shaped active site, but efficiently hydrolyzes longer maltooligosaccharides and soluble starch due to lower Km and higher kcat/Km for such substrates. To obtain structural insights into the mechanism governing its unique substrate specificity, a series of acarviosyl-maltooligosaccharides was employed for steady-state kinetic and structural analyses. The acarviosyl-maltooligosaccharides have a longer maltooligosaccharide moiety compared with the maltose moiety of acarbose, which is known to be the transition state analog of α-glycosidases. The clear correlation obtained between log Ki of the acarviosyl-maltooligosaccharides and log(Km/kcat) for hydrolysis of maltooligosaccharides suggests that the acarviosyl-maltooligosaccharides are transition state mimics. The crystal structure of the enzyme bound with acarviosyl-maltohexaose reveals that substrate binding at a distance from the active site is maintained largely by van der Waals interactions, with the four glucose residues at the reducing terminus of acarviosyl-maltohexaose retaining a left-handed single-helical conformation, as also observed in cycloamyloses and single helical V-amyloses. The kinetic behavior and structural features suggest that the subsite structure suitable for the stable conformation of amylose lowers the Km for long-chain substrates, which in turn is responsible for higher specificity of the longer substrates.

Solution Conformation of Konjac Glucomannan Single Helix

The possibility of the existence of konjac glucomannan (KGM) single helix in dilute solution was discussed finely by ultraviolet-visible spectrophotometer (UV-Vis) with Congo red complex, viscosimetry and circular dichroism (CD). KGM bound to Congo red and the λ max characteristic shifted markedly to longer wavelength (514 nm) in 0.25 M aq NaOH. The shifts of λ max and intrinsic viscosity were also compared in different mediums, such as different concentrations of KGM, NaOH, urea solutions or temperatures. The results indicated that KGM had an ordered, single-helical conformation in neutral, slight NaOH (<0.15 M) and urea (<3.0 M) solutions or at a moderate temperature (T<55ºС ), while it was transformed disorder, random coils in high NaOH (>0.45 M), urea (>4.0 M) solutions or at a high temperature (T>60ºС ). So it can be conclude that the outside factors play an important role on the stability of the single-helix conformation for KGM solution. The induced CD spectra by Congo red and the CD spectra of KGM in aqueous and urea solution indicated further that KGM had a single-helical structure. The new conclusion laid the foundation of determining helical radius, pitch and length of KGM in the future studies.

Development of synthetic double helical polymers and oligomers

AbstractThere is growing interest in the design and synthesis of artificial helical polymers and oligomers, in connection with biological importance as well as development of novel chiral materials. Since the discovery of the helical structure of isotactic polypropylene, a significant advancement has been achieved for synthetic polymers and oligomers with a single helical conformation for about half a century. In contrast, the chemistry of double helical counterparts is still premature. This short review highlights the recent advances in the synthesis, structures, and functions of double helical polymers and oligomers, featuring an important role of supramolecular chemistry in the design and synthesis of double helices. Although the artificial double helices reported to date are still limited in number, recent advancement of supramolecular chemistry provides plenty of structural motifs for new designs. Therefore, artificial double helices hold great promise as a new class of compounds. © 2009 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 47: 5195–5207, 2009

Double helix formation of poly(m-phenylene)s bearing achiral oligo(ethylene oxide) pendants and transformation into an excess of one-handed single helix through cholate binding in water

A water-soluble poly(m-phenylene) bearing an achiral oligo(ethylene oxide) chain at the 5-position was synthesized by the Ni(0)-mediated homo-coupling polycondensation of a 3,5-dibromophenol monomer. The poly(m-phenylene) adopted a single helical conformation in protic media and self-assembled into a double helix in water through aromatic interaction, while it took a random-coil conformation in chloroform. Upon the addition of sodium cholate in water, the double helical poly(m-phenylene) was transformed into single strands, which bound the cholate molecules to form an excess of one-handed single helix.

Characterization of dextrin hydrogels by FTIR spectroscopy and solid state NMR spectroscopy

Fourier transform infrared (FTIR) and 13C solid state nuclear magnetic resonance (NMR) spectroscopy were used to study dextrin structural changes occurring upon hydrogel formation by vinyl acrylate (VA) grafting and subsequent free radical polymerization. The degrees of VA substitution (DS) and polymerization (DP) were quantified up to 40%VA by FTIR intensity measurements and partial least squares (PLS)/FTIR, the latter being a faster and less error-prone method. Above 40%VA, both parameters are underestimated by FTIR. A spin counting NMR experiment showed high carbon observabilities for hydrogels and improved PLS/NMR models were achieved for DS and DP determination. Alternative NMR integration methods are hindered by the broad VA peaks and need for area correction, due to their CP dynamics. NMR changes in C1 profile showed that a single helical conformation predominates at lower %VA, being replaced by disordered conformations as %VA increases. Furthermore, a correlation FTIR/NMR study indicated that ring conformations are significantly affected in hydrogels, compared to unpolymerized dextrin.

Enhanced production of inducible nitric oxide synthase by beta-glucans in mice.

We have already demonstrated that various activities including NO (nitric oxide) synthesis in vivo and in vitro significantly differ between triple helical (SPG) and single helical (alkaline-treated SPG, SPG-OH) beta-glucans. It was previously suggested that the single helical conformer of beta-glucan (SPG-OH) was dominant in cytokine production and subsequent NO synthesis in vitro. In this study, we analyzed production of inducible nitric oxide synthase (iNOS) induced by beta-glucans in vitro and in vivo. The iNOS production was enhanced in proteose peptone-induced peritoneal macrophages (PMs) cultured with SPG-OH in the presence of IFN-gamma for 24 h, and SPG-OH-induced PMs. Moreover, SPG-OH was effective for iNOS production not only in isolated macrophages but also in tissue macrophages, whereas SPG was less effective. These findings suggest that a single helical conformer is essential for iNOS production, and that NO synthesis by beta-glucans is closely related to iNOS production.

Intramolecular Versus Intermolecular Induction of Helical Handedness in Pyridinedicarboxamide Oligomers

AbstractA new convergent synthetic scheme has been developed to prepare oligoamides of 2,6‐diaminopyridine and 4‐decyloxy‐2,6‐pyridinedicarboxylic acid. These compounds adopt stable single helical conformations in solution. NMR and circular dichroism studies show that intramolecular and intermolecular chiral induction of handedness in these helices is possible. Intramolecular induction is effected by attaching a single chiral group to the end of an oligomer. Intermolecular induction results from various noncovalent interactions between chiral carboxylic or sulfonic acids and the terminal residues of the oligoamides. Intramolecular induction appears to be more efficient than intermolecular induction. When both effects compete, intramolecular induction prevails. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)

Hydroxy-substituted oligopyridine dicarboxamide helical foldamers.

As shown by X-ray diffraction and NMR studies, short oligopyridine dicarboxamides bearing benzyloxy, hydroxy and hydroxylate moieties adopt very robust single helical conformations, even in water.

Helical molecular programming: folding of oligopyridine-dicarboxamides into molecular single helices.

Molecular strands composed of alternating 2,6-diaminopyridine and 2,6-pyridinedicarbonyl units have been designed to self-organize into single stranded helical structures upon forming intramolecular hydrogen bonds. Pentameric strands 11, 12, and 14, heptameric strands 1 and 20, and undecameric strand 15 have been synthesized using stepwise convergent strategies. Single helical conformations have been characterized in the solid state by single crystal X-ray diffraction analysis for four of these compounds. Helices from pentameric strands 12 and 14 extend over one turn, and helices from heptameric 20 and undecameric 15 species extend to one and a half and two and a half turns, respectively. Intramolecular hydrogen bonds are responsible for the strong bending of the strands. 1H NMR shifts both in polar and nonpolar organic solvents indicate intramolecular overlap between the peripheral aromatic groups. Thus, helical conformations also predominate in solution. Molecular stochastic dynamic simulations of strand folding starting from a high energy extended linear conformer show a rapid (600 ps at 300 K) conversion into a stable helical conformation.

Folding Dendrons: The Development of Solvent-, Temperature-, and Generation-Dependent Chiral Conformational Order in Intramolecularly Hydrogen-Bonded Dendrons

The synthesis of intramolecularly hydrogen-bonded dendrons with stereogenic terminal groups derived from (1S,2S)-(+)-thiomicamine up to the third generation is described. Circular dichroism (CD) studies reveal that the equilibria interconverting two diastereomeric helical conformations (M and P helices) relating a pair of anthranilamide termini depends on solvent, temperature, and dendrimer generation. A conformational preference for M-type helicity along the periphery of the dendrons increased with increasing dendrimer generation and in poor solvents as observed by CD. Equilibration of these diastereomeric helical conformations is rapid at the first generation in all solvents and at all temperatures investigated; however, at the second generation the equilibrium begins to bias a single diastereomeric helical conformation along the periphery that becomes maximal at low temperatures and in poor solvents. At the third generation, the helical bias is intrinsically higher so that the conformational preference...

Interconversion of single and double helices formed from synthetic molecular strands.

Synthetic single-helical conformations are quite common, but the formation of double helices based on recognition between the two constituent strands is relatively rare. Known examples include duplex formation through base-pair-specific hydrogen bonding and stacking, as found in nucleic acids and their analogues, and polypeptides composed of amino acids with alternating L and D configurations. Some synthetic polymers and self-assembled fibres have double-helical winding induced by van der Waals interactions. A third mode of non-covalent interaction, coordination of organic ligands to metal ions, can give rise to double, triple and quadruple helices, although in this case the assembly is driven by the coordination geometry of the metal and the structure of the ligands, rather than by direct inter-strand complementarity. Here we describe a family of oligomeric molecules with bent conformations, which exhibit dynamic exchange between single and double molecular helices in solution, through spiral sliding of the synthetic oligomer strands. The bent conformations leading to the helical shape of the molecules result from intramolecular hydrogen bonding within 2'-pyridyl-2-pyridinecarboxamide units, with extensive intermolecular aromatic stacking stabilizing the double-stranded helices that form through dimerization.

Interleukin 8 production of human leukocytes stimulated by triple or single helical conformer of an antitumor (1?3)-?-D-glucan preparation, sonifilan

Previous research has suggested that the immunopharmacological activity of soluble (1→3)-β-D-glucan in mice depends on its conformation. In the present study, we examined the relationship between conformation of Sonifilan (SPG), a high molecular weight (1→3)-β-D-glucan, and interleukin 8 (IL-8) productivity in an in vitro human system. All experiments were performed using a polypropylene tube in order to reduce cellular adherence and reduce integrin-mediated signaling. SPG takes the form of a triple helical conformer in water, which changes in response to treatment with an aqueous sodium hydroxide to a single helical conformer (SPG-OH). Whole blood, peripheral blood mononuclear cells (PBMC), and neutrophils (PMN) were stimulated with either SPG or SPG-OH and then cultured in vitro. Next, IL-8 production of the cells and supernatants was measured immunochemically (ELISA and FACS). In whole blood and PBMC cultures, SPG-OH induced higher IL-8 production than SPG. In PMN culture, IL-8 production induced by SPG was comparable to that of SPG-OH. Enhanced IL-8 production of monocytes and PMN was confirmed by detection of increased levels of intracellular IL-8 cultured in the presence of blefeldin and then assessed by FACS. These findings suggested that SPG and SPG-OH enhanced the productivity of IL-8 in human leukocytes.

Interleukin 8 production of human leukocytes stimulated by triple or single helical conformer of an antitumor (1→3)‐β‐D‐glucan preparation, sonifilan

Interleukin 8 production of human leukocytes stimulated by triple or single helical conformer of an antitumor (1→3)‐β‐D‐glucan preparation, sonifilan

Enzyme immunoassay system for estimating the ultrastructure of (1,6)-branched (1,3)-β-glucans

A sandwich-type enzyme immunoassay (EIA) system for quantifying branched (1,3)-β-glucans was established. A polyclonal antibody was purified with antigen-conjugated aminocellulofine and labelled with biotin to be used as the detection antibody. The reactivity of the antibody was restricted to only (1,6)-branched (1,3)-β-glucans. Molecular weight dependency of (1,6)-branched (1,3)-β-glucan in the reactivity was also observed. Alkaline-treated (1,6)-branched (1,3)-β-glucan which was reported to be a single helical conformer, showed a lower absorbance compared to the untreated triple helix conformer. The conformational alteration of the single helix to the triple helix was produced by heating for 15 min at 100°C. The results suggest that EIA has higher reactivity to the triple helical ultrastructure of (1,6)-branched (1,3)-β-glucans, and can be applied to estimate the conformational changes of (1,6)-branched (1,3)-β-glucans.

Physical properties of xanthan, galactomannan and their mixtures in aqueous solutions

AbstractSome new information on the conformation of xanthan in aqueous solutions is given. A single helical chain conformation characterizes xanthan in the native state. When heated over the critical temperature for conformational change ( helixŕ coil), the xanthan is denatured and renatured when it is cooled down in a localy double helical structure. A galactomannan was also characterized and its persistence length was obtained (Lp ≈︁ 90A°). Then mixtures of the galactomannan and xanthan were investigated to propose a mechanism for the specific gelation. From the results of microcalorimetry and circular dichroism, it is concluded that a complex is formed between one disordered xanthan chain and one galactomannan chain and that an ordered conformation is stabilized at temperatures lower than 25°C when the galactomannan has a M/G ratio of ≈︁ 3. This temperature corresponds to the sol‐gel transition. This is the first time that a structure of the crosslink points is demonstrated.