PNA‐Encoded Synthesis (PES) of a 10 000‐Member Hetero‐Glycoconjugate Library and Microarray Analysis of Diverse Lectins

Identification of selective and synthetically tractable ligands to glycan‐binding proteins is important in glycoscience. Carbohydrate arrays have had a tremendous impact on profiling glycan‐binding proteins and as analytical tools. We report a highly miniaturized synthetic format to access nucleic‐acid‐encoded hetero‐glycoconjugate libraries with an unprecedented diversity in the combinations of glycans, linkers, and capping groups. Novel information about plant and bacterial lectin specificity was obtained by microarray profiling, and we show that a ligand identified on the array can be converted to a high‐affinity soluble ligand by straightforward chemistry.     

Cellular Antisense Activity of PNA–Oligo(bicycloguanidinium) Conjugates Forming Self‐Assembled Nanoaggregates

A series of peptide nucleic acid–oligo(bicycloguanidinium) (PNA–BG_n) conjugates were synthesized and characterized in terms of cellular antisense activity by using the pLuc750HeLa cell splice correction assay. PNA–BG₄ conjugates exhibited low micromolar antisense activity, and their cellular activity required the presence of a hydrophobic silyl terminal protecting group on the oligo(BG) ligand and a minimum of four guanidinium units. Surprisingly, a nonlinear dose–response with an activity threshold around 3–4 μM , indicative of large cooperativity, was observed. Supported by light scattering and electron microscopy analyses, we propose that the activity, and thus cellular delivery, of these lipo‐PNA–BG₄ conjugates is dependent on self‐assembled nanoaggregates. Finally, cellular activity was enhanced by the presence of serum. Therefore we conclude that the lipo‐BG‐PNA conjugates exhibit an unexpected mechanism for cell delivery and are of interest for further in vivo studies.     

Inhibitor and protein microarrays for activity-based recognition of lipolytic enzymes

Protein and small-molecule microarrays are useful tools for high-throughput analysis of DNA-protein, protein-protein, and protein-small molecule interactions. Here we report on novel microarrays for activity screening of lipases and esterases based on phosphonic acid ester inhibitors. These compounds are activity recognition probes (ARPs) and bind to active serine hydrolases in a stoichiometric and irreversible manner. Protein microarrays were generated by spotting six different lipolytic enzymes onto hydrogel-coated glass slides. The activity of immobilized enzymes was determined after treatment with fluorescently labeled ARPs. Alternatively, biotinylated ARPs were bound to streptavidin slides in order to identify their affinity for enzymes in solution. Both systems, the protein- and ARP microarrays proved to be useful and versatile tools for the rapid identification and characterization of novel and known lipolytic enzymes.     

Structural insights into peptide self-assembly using photo-induced crosslinking experiments and discontinuous molecular dynamics

Determining the structure of the (oligomeric) intermediates that form during the self-assembly of amyloidogenic peptides is challenging because of their heterogeneous and dynamic nature. Thus, there is need for methodology to analyze the underlying molecular structure of these transient species. In this work, a combination of fluorescence quenching, photo-induced crosslinking (PIC) and molecular dynamics simulation was used to study the assembly of a synthetic amyloid-forming peptide, Aβ_16-22. A PIC amino acid containing a trifluormethyldiazirine (TFMD) group—Fmoc(TFMD)Phe—was incorporated into the sequence (Aβ*_16–22). Electrospray ionization ion-mobility spectrometry mass-spectrometry (ESI-IMS-MS) analysis of the PIC products confirmed that Aβ*_16–22 forms assemblies with the monomers arranged as anti-parallel, in-register β-strands at all time points during the aggregation assay. The assembly process was also monitored separately using fluorescence quenching to profile the fibril assembly reaction. The molecular picture resulting from discontinuous molecule dynamics simulations showed that Aβ_16-22 assembles through a single-step nucleation into a β-sheet fibril in agreement with these experimental observations. This study provides detailed structural insights into the Aβ_16-22 self-assembly processes, paving the way to explore the self-assembly mechanism of larger, more complex peptides, including those whose aggregation is responsible for human disease.     

Polysaccharide microarrays for polysaccharide-platelet-derived-growth-factor interaction studies

Polysaccharide microarrays have great potential for the high-throughput analysis of polysaccharide-protein interactions. Here we demonstrate that a polysaccharide microarray prepared by printing a library of dextran polymers derivatized by methylcarboxylate, benzylamide, and sulfate groups (DMCBSu compounds) on to glass slides permitted the rapid identification of a set of compounds able to interact with the platelet-derived growth factor BB (PDGF-BB) isoform, a growth factor involved in wound healing. Microarray interaction results were compared to the capacity of DMCBSu compounds to potentiate the in vitro PDGF-BB-induced proliferation of human dermal fibroblasts.     

‘Bricks and mortar’ nanoparticle self‐assembly using polymers

Developments in self‐assembly methods allow access to hierarchical materials featuring a wide range of functionality and applications. Polymer‐based self‐assembly of nanoparticles opens up new avenues for the fabrication of highly structured nanocomposites that can serve as bridges between ‘bottom‐up’ and ‘top‐down’ methods. Of various interactions leading to self‐assembly of nanocomposites, hydrogen bonding and electrostatic interactions are commonly utilized. In this review, we illustrate the design and subsequent property tuning of various self‐assembled nanocomposite materials that were developed based on these interactions. Copyright © 2007 Society of Chemical Industry     

Alternative procedure for the incorporation of quantum dots into poly(N‐isopropyl acrylamide‐co‐acrylic acid) microgels based on multiple interactions

Monodisperse fluorescent poly(N‐isopropyl acrylamide‐co‐acrylic acid) microgels doped with quantum dots (QDs) were fabricated as follows. First, cysteamine‐capped cadmium telluride (CA–CdTe) QDs were introduced into the microgels at pH 7 by electrostatic interactions. Afterward, the CA–CdTe QDs were further immobilized in the microgels by the collapse of the polymer network when the pH of solution was adjusted to 4. In this system, there existed multiple interactions between the CA–CdTe QDs and the microgels, including hydrogen bonds, electrostatic interactions, and coordination bonds. The photoluminescence intensity and maximum emission wavelength of the resulting microgels could be easily adjusted by changes in the content of the CA–CdTe QDs in the hybrid microgels (HMs) and with differently sized QDs, respectively. We found that the lower the addition of CA–CdTe QDs was, the bigger the blueshift of the photoluminescence spectra of the HMs was and the weaker the photoluminescence intensity was. Finally, temperature‐responsive emission of the HMs was examined. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2016 , 133, 43227.     

Insights into Carbopol gel formulations: Microscopy analysis of the microstructure and the influence of polyol additives

Carbopol is a family of high molecular weight cross‐linked poyacrylic acids widely used as thickening agents in several industrial applications. In this work, the structure of Carbopol 974P NF was investigated at pH 7.4 by means of capillary viscometry, dynamic rheology, fluorescence microscopy, and electron microscopy in the dilute and semi‐dilute (hydrogel) state. Especially, high‐pressure freezing technique has proved to be a technique of choice to visualize the Carbopol hydrogel microstructure through cryo‐scanning electron microscopy analysis without artifacts related to the formation of ice crystals. It has been shown that the Carbopol hydrogel has a hierarchical structure made of large spherical entities of a few micrometers in size packed together and composed of much smaller agglomerated microgel particles. In addition, fluorescence and electron microscopies have pointed out that the presence of polyol additives in the gel formulation favors the formation of more dense or jammed structure, probably through hydrogen bonding. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42761.     

Analyzing Xyloglucan Endotransglycosylases by Incorporating Synthetic Oligosaccharides into Plant Cell Walls

The plant cell wall is a cellular exoskeleton consisting predominantly of a complex polysaccharide network that defines the shape of cells. During growth, this network can be loosened through the action of xyloglucan endotransglycosylases (XETs), glycoside hydrolases that “cut and paste” xyloglucan polysaccharides through a transglycosylation process. We have analyzed cohorts of XETs in different plant species to evaluate the substrate specificities of xyloglucan acceptors by using a set of synthetic oligosaccharides obtained by automated glycan assembly. The ability of XETs to incorporate the oligosaccharides into polysaccharides printed as microarrays and into stem sections of Arabidopsis thaliana, beans, and peas was assessed. We found that single xylose substitutions are sufficient for transfer, and xylosylation of the terminal glucose residue is not required by XETs, independent of plant species. To obtain information on the potential xylosylation pattern of the natural acceptor of XETs, that is, the nonreducing end of xyloglucan, we further tested the activity of xyloglucan xylosyl transferase (XXT) 2 on the synthetic xyloglucan oligosaccharides. These data shed light on inconsistencies between previous studies towards determining the acceptor substrate specificities of XETs and have important implications for further understanding plant cell wall polysaccharide synthesis and remodeling.     

Studies on self‐assembly and characterization of polyelectrolytes and organic dyes

Four polyelectrolyte complexes were formed through the self‐assembly of poly‐N‐ethyl‐N,N‐ dimethylamino ethyl methacrylate (PEDEM) and poly‐ N‐ethyl‐4‐vinylpyridinium (PEVP) cations with methyl orange (MO) and metanil yellow (MY) anions in water. The FTIR spectra showed that the assembly was formed chiefly through electrostatic force and hydrophobic interaction between polyelectrolytes and organic dyes without new bonds emerging. The fluorescence spectra revealed that the emission waves of the complexes of PEDEM‐MY and PEDEM‐MO in alcohol were blue‐shift in comparison with those of dyes in alcohol, and the emission waves of the PEVP‐MY and PEVP‐MO complexes in alcohol were red‐shift in comparison with those of dyes in alcohol. The structure of the complexes in solid state were also investigated by differential thermal analysis (DTA) and X‐ray diffraction experiments. It was proved that the complexes were new materials formed through weak interactions. © 2002 Wiley Periodicals, Inc. J Appl Polym Sci 85: 638–644, 2002     

Effects of monovalent and multivalent ions on the stability of a polyelectrolyte complex with entrapped surfactants

In this article, we report the effects of nine different monovalent and multivalent salts on the particle size and stability of refined, positively charged polyelectrolyte complexes (PECs) with entrapped surfactant. Dynamic light scattering and ζ potential measurements of these polymeric particles as a function of various salt concentrations showed that both counter ions and co‐ions induced a concentration‐dependent increase in the particle size and a decrease in the ζ potential. We found that the anion concentration where the particle size doubled and the maximum anion concentration beyond which particle precipitation occurred (C_a,max) demonstrated a power law dependence on the anion valence. Moreover, for anions of the same valence but different hydration radio, C_a,max decreased in the following order: NO₃ → Cl → HPO₄^2? > SO₄^2? > PO₄^3?. However, unlike the case of hard colloids where co‐ions have relatively little effect on particle interactions, the co‐ions also increased the hydrodynamic radii of our PECs in the following order: K⁺ ≈ Na⁺ > Ca²⁺ > Mg²⁺ > Al³⁺. Furthermore, we found that the entrapped surfactants were shielded from the adverse effect of multivalent ions; this established that the monovalent and multivalent ions interacted with the polyelectrolyte shells of the PEC. This behavior was in contrast with the effect of salts on mixtures containing the polyelectrolyte and surfactant components, where the addition of salts typically causes an interaction with the individual components. Because several biomedical and technological applications involving PECs require saline environments, our studies provide insight into how small ions influence the PEC stability in applications involving varying salinities. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42099.     

Self‐assembly of rod‐coil molecules into lateral chain‐length‐dependent supramolecular organization

In this article, we report synthesis and characterization of the self‐assembly behavior of coil‐ rod‐coil molecules, consisting of four biphenyls and a p‐terphenyl unit linked together with ether bonds as a rod segment. These molecules contain lateral methyl or ethoxymethyl groups at 2 and 5 positions of the middle benzene ring of p‐terphenyl. The self‐assembling behavior of these materials was investigated by means of DSC, POM, and SAXS in the bulk state. The results reveal that self‐assembling behavior of these molecules is dramatically influenced by a lateral methyl or ethoxymethyl groups in the middle of rod segment. In addition, molecule with PEO (DP = 17) coil chains of identical coil volume fraction to the corresponding molecule connected by PPO (DP = 12) coil chains, shows diverse self‐organizing behavior that may result from the parameters of cross‐sectional area of coil segment and the steric hindrance at the rod/coil interface. © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2011     

Redox‐active LBL films via in situ template polymerization of hydroquinone

New redox‐active hydrogel thin films with poly(hydroquinone) (PHQ) as polymeric redox couple and chitosan hydrogel as matrix are synthesized. Single‐component layer‐by‐layer films of chitosan are first fabricated in a uniform and reproducible manner, which is proved by UV and AFM studies. The chitosan films are then treated in acidic hydroquinone (HQ) solution to introduce PHQ. In situ generation of PHQ in the films is proved by UV and FTIR spectroscopy. PHQ formation also results in an increased film roughness and thickness. We confirmed that oxygen acts as oxidant and chitosan acts as template in the polymerization of HQ. Cyclic voltammogram of the film presents two oxidation peaks at 0.21 and 0.75 V and one reduction peak at ?0.15 V(vs Ag/AgCl), indicating the film, which is originally not redox‐active, becomes redox‐active after the introduction of PHQ. The new redox‐active films may find applications in amperometric biosensors. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013     

Studies on self‐assembly and characterization of polyelectrolytes and organic dyes

Four polyelectrolyte complexes were formed through self‐assembly of poly‐N‐ethyl‐N,N‐(dimethylamino)ethyl methacrylate (PEDEM) and poly‐N‐ethyl‐4‐vinylpyridinium (PEVP) cations with methyl orange (MO) and metanil yellow (MY) anions in water. The FTIR spectra showed that the assembly was formed chiefly through electrostatic force and hydrophobic interaction between polyelectrolytes and organic dyes without new bonds emerging. The fluorescence spectra revealed that the emission waves of the complexes of PEDEM–MY and PEDEM–MO in alcohol were blue‐shifted in comparison with that of dyes in alcohol, and the emission waves of the PEVP–MY and PEVP–MO_ complexes in alcohol were red‐shifted in comparison with that of dyes in alcohol. The structure of the complexes in the solid state were also investigated by DTA and X‐ray diffraction experiments. It could be proved that the complexes were new materials formed through weak interactions. © 2002 Wiley Periodicals, Inc. J Appl Polym Sci 87: 369–374, 2003