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Highlights: [•] Saccharidic chiral dopants for nematic liquid crystals were synthesized from d-glucose. [•] The helical twisting power (HTP) values were evaluated. [•] The chiral dopants induced helices in the host nematic liquid crystals. [•] No relationship between HTP and specific rotation on the chiral dopants was observed in this study. [Copyright &y& Elsevier]

Abstract: Conversions of the C-5 acetamide group in sialic acid into two kinds of Cbond substituents were accomplished under Shotten–Baumann conditions. The polymerizable glycomonomers also contain a hydrophobic chain or hydroxyl group at the anomeric position. Radical polymerizations of the fully protected glycomonomers were carried out with acryl amide in the presence of ammonium persulfate (APS) and N,N,N′,N′-tetramethylethylenediamine (TEMED), followed by deprotection to furnish water-soluble glycopolymers. The activities of the deprotected glycopolymers and glycomonomers against human influenza viruses (H1N1 and H3N2) and avian influenza virus (H5N3) were evaluated. Biological evaluations showed that the glycomonomers containing a long hydrophobic chain at the anomeric position had both hemagglutination and neuraminidase inhibitory activities. [Copyright &y& Elsevier]

Abstract: A series of bi-fluorescence-labeled maltooligosaccharides that lead to fluorescence resonance energy transfer (FRET) was systematically synthesized. Effective FRETs were observed with all of the synthesized probes. Digestion of probes having tetra-, quintet-, hexa- or hepta-saccharidic chain lengths with human saliva α-amylase resulted in disappearance of FRET when an excitation wavelength of at 290nm was used followed by detection at ca. 520nm due to emission from the dansyl moiety. However, continuous FRET was observed when probes having di- or trisaccharidic chain lengths were used as substrates. In addition to the substrate characteristics based on saccharidic chain length, the reaction rates of digestion for the substrates by amylase were different and also depended on their saccharidic chain length. [Copyright &y& Elsevier]

Abstract: In order to develop novel influenza sialidase inhibitors, we constructed a library of glycoclusters composed of twelve types of sialylated dendrimers with thioglycosidic linkage that are resistant to hydrolysis by the sialidases. These sialodendrimers were synthesized by condensation reaction between a thiosialoside modified on the aglycon terminal end by a thioacetyl group and twelve types of carbosilane dendrimers having brominated terminal ends under deacetylation conditions, and temporal re-protection was performed for purification. Removal of all protection of the glycodendrimers was accomplished by transesterification and subsequent saponification to provide corresponding water-soluble glycodendrimers in good yields. For investigation of the structure–activity relationship, dendrimer scaffolds having differences in number of the sugar moieties, such as 3-, 4-, 6- and 12-functionalized dendrimers, and in linkage patterns, such as normal aliphatic linkage, ether- and amide-linkages. Biological evaluations of these glycodendrimers showed that all of the ether- and amide-elongated compounds had inhibitory potencies for the influenza sialidases in the mM range, while compounds having normal aliphatic linkage did not have any activities except for a 12-functionalized compound. [Copyright &y& Elsevier]

The thioglycoside of Neu5Gc unit was synthesized from the Neu5Ac derivative in 4 steps. The thioglycoside of Neu5Gc as a glycosyl donor was allowed to react with various alcohols to afford the corresponding glycosides. [Display omitted] • A Neu5Gc unit was prepared from a Neu5Ac derivative. • Thioglycoside of Neu5Gc was synthesized as a glycosyl donor. • Glycosidation of the Neu5Gc derivative gave the desired Neu5Gc-related compound. • Reaction properties of the thioglycoside of Neu5Gc were investigated. Efficient synthesis of lauryl thioglycoside of Neu5Gc was accomplished from a known thioglycoside of Neu5Ac. The reactivities of the Neu5Gc thioglycoside for O -glycosidation by means of TMSOTf-NIS as a promoter system were investigated, and the thioglycoside showed good activities with simple alcohols. In addition to the simple alcohols, the reactivities of lauryl thioglycoside of Neu5Gc were also evaluated with glucose and lactose derivatives and the results suggested that the thioglycoside is a potential candidate for glycosidation as the glycosyl donor. [ABSTRACT FROM AUTHOR]

Abstract: An efficient synthesis of a series of carbosilane dendrimers uniformly functionalized with α-thioglycoside of sialic acid was accomplished. The results of a preliminary study on biological responses against influenza virus sialidases using thiosialoside clusters showed that some of the glycodendrimers have inhibitory potencies against the sialidases. [Copyright &y& Elsevier]

• Two polymers having α-mannoside moieties or dansyl fluorophores were conventionally prepared. • Efficient synthesis of glycopolymers having α-mannoside moieties and the fluorophores was accomplished. • Biological evaluations of glycopolymers—lectin interactions were performed. • Intermolecular FRET was clearly observed when fluorogenic glycopolymers were used. Quantitative evaluations of the binding affinities between glycopolymers and lectins in order to investigate the functionality of glycopolymers are of great importance. In this study, we developed fluorogenic glycopolymers that induce Förster resonance energy transfer (FRET) upon binding to lectins and we attempted directly to monitor the interaction followed by evaluation of the binding affinity on the basis of intermolecular FRET. Water-soluble glycopolymers with various polymer compositions containing mannose moieties as ligands for a lectin and dansyl moieties as fluorogenic chromophores for the FRET system were prepared, and the binding constant K a (M−1) with concanavalin A (Con A; mannose-binding lectin) was estimated on the basis of the change in fluorescence intensity caused by FRET. Fluorescence measurements revealed intermolecular FRET between the fluorogenic glycopolymer and Con A, and the binding constants were estimated to be 0.398×106 ∼ 2.40×106 (M−1). The results suggested that this method can be applied to evaluation of the interaction when the binding of a glycopolymer to a lectin results in little fluorescence change. [Display omitted] [ABSTRACT FROM AUTHOR]

Abstract: Synthesis of bi-fluorescence-labeled maltooligosaccharides for amylase assay was accomplished. Preliminary biological evaluation of both bi-fluorescence-labeled maltohexasaccharide and maltose using α-amylase was carried out, and the hexaosyl derivative showed unique variation on the basis of fluorescence resonance energy transfer (FRET). [Copyright &y& Elsevier]

[Display omitted] • Homopolymers and copolymers with histidine COOH, CONH 2 , and NH 2 were synthesized. • The polymers were incubated with bovine serum albumin and analyzed by SDS-PAGE. • Polymers with histidine COOH cleaved bovine serum albumin at pH 4.2 and pH 7.4. • Several polymers with specific histidine densities showed activity at pH 8.0. • Polymers with specific COOH density exhibited varying digestion patterns based on pH. Artificial enzymes with serine, aspartic acid, and histidine that mimic the catalytic triad of serine proteases have been studied. However, they were mainly investigated for their cleavage activity on low molecular-weight esters and amides, and there has been a lack of knowledge about their protein cleavage activity. We previously found that not only ternary polyacrylamides with serine, aspartic acid, and histidine but also homo-polyacrylamides with a single type of amino acid cleaved bovine and human serum albumins. These results suggested that the carboxy group, a common functional group in active polymers, may be involved in the expression of activity. In this study, we synthesized histidine polymers in which the carboxy group was changed to a carboxamido or amino group and investigated the relationships between structure and activity. Histidine homopolymers and histidine-acrylamide copolymers were incubated with bovine serum albumin at 37 °C for one week under different pH conditions and then analyzed by sodium dodecyl-sulfate polyacrylamide gel electrophoresis. In the case of histidine homopolymers, the carboxy-type homopolymers were active under weakly acidic to neutral conditions, while the carboxamido- and amino-type homopolymers were not active regardless of liquid properties. In the case of histidine-acrylamide copolymers, their activity was essentially the same as that of each type of histidine homopolymer, but for all types of copolymers, activity appeared under weakly basic conditions when the histidine monomer units were of a specific density. The carboxy-type copolymers with dual-mode activity showed different degradation patterns of bovine serum albumin under acidic and basic conditions. The structural features common to the active homopolymers and copolymers suggested that the carboxy group contributes to the activity under weakly acidic to neutral conditions, while the imidazole group of the histidine side chain and its density contribute to the activity under weakly basic conditions. [ABSTRACT FROM AUTHOR]

[Display omitted] • Fischer's glycoside synthesis effectively worked with alcohols with longer aliphatic chains. • Glycopolymers having α-glycoside-type GlcNAc moieties were prepared. • Sugar densities on the glycopolymers were controlled. • Biological evaluations of glycopolymers—WGA were tested. • Glycopolymer having an appropriate sugar-density showed the highest affinity. Fischer's glycoside synthesis was applied to linker precursor alcohols of two different lengths having appropriate alkane chains to obtain the corresponding α-glycoside and it was found to be applicable with moderate yields. Water-soluble glycomonomers were systematically prepared from N -acetyl- d -glucosamine (GlcNAc) by introducing two kinds of alcohols having different methylene lengths. Typical radical polymerizations of the glycomonomers with acrylamide as a modulator for control of the distance between carbohydrate residues in water in the presence of ammonium persulfate (APS)- N , N , N' , N' -tetramethylethylenediamine (TEMED) gave a series of glycopolymers with various α-glycoside-type GlcNAc residue densities. Fluorometric analysis of the interaction of wheat germ agglutinin (WGA) with the glycopolymers was performed and the results showed unique binding specificities based on structural differences. [ABSTRACT FROM AUTHOR]

Bivalent glycoconjugates have a minimal valence with avidity potential on protein-carbohydrate interactions as well as simplicity of chemical structures enabling simple synthesis with low cost. Understanding the way to maximize the affinities of bivalent glycoconjugates is important for the development of cost-effective tools for therapeutic and diagnostic research. However, there has been little discussion about the effects of constraints imposed from ligand scaffolds on the binding abilities. We synthesized three kinds of biantennary N -acetylglucosamine glycosides with different scaffolds using isobutenyl bis(propargyl)ether as a common scaffold precursor. Decoration of the scaffold branches with GlcNAc moieties through copper-catalyzed azide-alkyne cycloaddition and grafting of the alkenyl focal point to another bivalent biotin dendron through thiol-ene and nucleophilic substitution reactions were successfully carried out in an orthogonal manner. The association constants of the ligands against wheat germ agglutinin were determined by a fluorometric titration assay. A bivalent biotin counterpart provided higher affinity than an isobutyl scaffold, whereas an isobutenyl scaffold yielded more enhancement than a bivalent biotin counterpart. The present work suggested that the constraint and steric bulk of ligand scaffolds are possible factors for improving binding properties of glycoconjugates against lectins or proteins. [ABSTRACT FROM AUTHOR]

Survivin, an inhibitor of the apoptosis protein family, is a potent tumor marker for diagnosis and prognosis. The enzyme-linked immunosorbent assay (ELISA) is one of the methods that has been used for detection of survivin. However, ELISA has several disadvantages caused by the use of conventional antibodies, and we have therefore been trying to develop a novel ELISA system using camelid single-domain antibodies (VHHs) as advantageous replacements. Here we report a supplemental approach to improve the VHH-polyclonal antibody sandwich ELISA for survivin detection. Iodoacetyl-functionalized pullulan was synthesized, and its thiol reactivity was characterized by a model reaction with l -cysteine. The thiophilic pullulan was applied to an immunoassay as an additive upon coating of standard assay plates with an anti-survivin VHH fusion protein with C -terminal cysteine. The results showed that the mole ratio of the additive to VHH had a significant effect on the consequent response. Mole ratios of 0.07, 0.7, and 7 led to 90% lower, 15% higher, and 69% lower responses, respectively, than the response of a positive control in which no additive was used. The background levels observed in any additive conditions were as low as that of a negative control lacking both VHH and the additive. These results indicate the applicability of the thiol-reactive pullulan as a response enhancer to VHH-based ELISA. [ABSTRACT FROM AUTHOR]

[Display omitted] Glycopolymers having sialyl α2-3 lactose moieties via longer spacer arms were systematically prepared from the corresponding glycomonomers. Radical polymerization of glycomonomers gave a series of glycopolymers displaying various sugar densities. Fluorometric analyses of wheat germ agglutinin (WGA) against the glycopolymers were conducted and the results showed unique binding specificities on the basis of sugar densities. [ABSTRACT FROM AUTHOR]

We assessed the relationship between fractional anisotropy (FA) values of magnetic resonance-diffusion tensor imaging (DTI) and long-term outcome in patients with hemiparesis after intracerebral hemorrhage (N =16). DTI data were obtained on days 14–18. FA values within the cerebral peduncle were analyzed using a computer-automated method. Motor outcome of hemiparesis was evaluated using Brunnstrom stage (six-point scale: severe to normal) for separate shoulder/elbow/forearm, wrist/hand, and lower extremity functions when patients were discharged from a long-term rehabilitation facility 2–7months after onset. In addition, the motor component of the functional independence measure (FIM-motor) was scored. The ratio of FA values in the affected hemisphere to those in the unaffected hemisphere (rFA) was assessed in relation to the clinical data (Spearman’s rank correlation test, P <0.05). Analysis revealed a statistically significant relationship between rFA and upper extremity function (R =0.841 for shoulder/elbow/forearm; 0.855 for wrist/hand). Although statistically significant, the relationship between rFA and lower extremity function (R =0.713) was less evident. The relationship between rFA and lower extremity and FIM-motor scores was moderate (R =0.579). These findings suggest that DTI is a potentially useful tool for outcome prediction, especially for upper extremity function. [Copyright &y& Elsevier]

[Display omitted] • Glycopolymer having GlcNAc moieties was prepared. • Sugar densities on the glycopolymers were controlled. • Degree of freedom of GlcNAc residue on the glycopolymers was controlled. • Biological evaluations of glycopolymers—lectin were performed. Four kinds of water-soluble carbohydrate monomers having different methylene numbers of polymerizable aglycons were systematically prepared from N -acetyl-D-glucosamine (GlcNAc). Typical radical polymerization in the presence of ammonium persulfate (APS)—tetramethylethylenediamine (TEMED) in water was applied for the monomers with acryl amide as a regulator for adjusting the average distance between carbohydrate residues, a series of glycopolymers having various densities of GlcNAc residues was obtained. Regulation of the polymers showed two pivotal points: one is the effect of lengths between carbohydrate residues and the polymer backbone and the other one is the influence of average densities of carbohydrates in the glycopolymer. Carbohydrate—protein interaction by means of the glycopolymer library for wheat germ agglutinin (WGA) as a model protein was investigated and the highest K a value was obtained when the glycopolymer having the longest methylene length and the ratio of 1:10 (sugar unit:acrylamide unit) was used as a substrate. [ABSTRACT FROM AUTHOR]

Highlights: [•] Chloromethylstyrene and N-acetyl-d-glucosamine (GlcNAc) were used for this study. [•] Simple modification of the starting materials gave the water-soluble glycomonomer. [•] A convenient access for glycopolymers was accomplished by radical polymerization. [•] The glycopolymers exhibited high affinity against WGA. [ABSTRACT FROM AUTHOR]

[Display omitted] • A hydrophobic Pba and a hydrophilic glycomonomer were polymerized. • The polymerization gave the corresponding water-soluble glycopolymer having Pba moieties. • Structural elucidation of the polymer was performed by 1H NMR. • The thermal property of the polymers was evaluated on the basis of TG-DTA measurement. • Uv–vis spectroscopic analysis of the polymer showed the characteristic absorption due to Pba. The first trial of incorporation of pheophorbide a (Pba) into a synthetic glycopolymer giving water-soluble ability was carried out, and the polymerization reaction of each monomer proceeded in DMSO to afford the desired glycopolymer having Pba side chains. Characterization of the polymer was performed by means of spectroscopic analyses and thermal studies, and the results supported the desired macromolecular architecture. [ABSTRACT FROM AUTHOR]

Abstract: An efficient synthesis of sialyllactosamine (SiaLacNAc) clusters using carbosilanes as core scaffolds has been accomplished by means of chemical and enzymatic approaches. N-Acetyl-d-glucosamine (GlcNAc) clusters having O-glycosidic linkage or S-glycosidic linkage were chemically synthesized from known intermediates in high yields. The GlcNAc clusters were first used as substrates for β1,4 galactosyl transferase using UDP-galactose (UDP-Gal) as a sugar source to provide corresponding N-acetyllactosamine clusters. Further sugar elongation of the LacNAc clusters was demonstrated using α2,3 sialyl transferase and CMP-neuraminic acid (CMP-NANA) to yield the corresponding SiaLacNAc clusters. [ABSTRACT FROM AUTHOR]

Abstract: Synthetic construction of fucosyl chitobiose [GlcNAcβ1→4(Fucα1→3)GlcNAc] as an allergy-associated carbohydrate epitope was accomplished from three building blocks. The trisaccharidic unit was further transformed into a carbohydrate monomer and polymerization of the glycomonomer proceeded smoothly to provide a series of glycopolymers having various carbohydrate densities. In addition to the organic syntheses, biological evaluations of the glycomonomer and the polymers were carried out and sugar-clustering effects were observed. [Copyright &y& Elsevier]

Abstract: Glycomonomers of sialic acid in which the acetamide group at C-5 was converted into two kinds of Cbond substituents were prepared and the fully protected glycomonomers were directly polymerized before deprotection steps. Radical polymerization with acrylamide in DMF in the presence of ammonium persulfate and N,N,N’,N’-tetramethylethylenediamine proceeded smoothly and gave corresponding sialopolymers. Interestingly glycomonomers had hemagglutination inhibitory activities not only for H1N1 but also for H3N2 of human influenza virus strains. [Copyright &y& Elsevier]

Abstract: A series of carbosilane dendrimers uniformly functionalized with sialyl lactose moieties (Neu5Acα2→3Galβ1→4Glc) was systematically synthesized, and biological evaluations for anti-influenza virus activity using the glycodendrimers were performed. The results suggested that the glycodendrimers had unique biological activities depending on the form of their core frame, and Dumbbell(1)6-amide type glycodendrimer 7 showed particularly strong inhibitory activities against human influenza viruses [A/PR/8/34 (H1N1) and A/Aichi/2/68 (H3N2)]. The results suggested that the structure–activity relationship (SAR) on the glycolibrary against various influenza viruses was observed, and dumbbell-shaped dendrimers as supporting carbohydrate moieties were found to be the most suitable core scaffolds in this study. [Copyright &y& Elsevier]

Abstract: An efficient separation between fully acetylated thiosialoside methyl esters and fully acetylated Neu5Ac2en methyl esters was accomplished by means of a size-exclusion chromatography (SEC) method. Purity determinations and structural elucidation of the isolated compounds were performed by a combination of elemental analyses and spectroscopic analyses, including IR, 1H, and 13C NMR, and mass spectroscopic analyses. [Copyright &y& Elsevier]

Abstract: An efficient synthesis of a series of carbosilane dendrimers uniformly functionalized with sialyl α(2→3) lactose (Neu5Acα(2→3)Galβ(1→4)Glcβ1→) moieties was accomplished. The results of a preliminary study on biological responses against influenza virus hemagglutinin, using the sialyl lactose clusters showed unique biological activities on the basis of the structure–activity relationship according to the carbosilane scaffolds. [Copyright &y& Elsevier]

Abstract: A conventional synthesis of α-thioglycoside of sialic acid as a glycomonomer was accomplished. Radical copolymerization of the glycomonomer with vinyl acetate proceeded smoothly to afford a new class of glycopolymers having thiosialoside residues, in which all protection was removed by a combination of transesterification and saponification to provide a water-soluble thiosialoside cluster. The results of a preliminary study on biological responses against influenza virus neuraminidases using the thiosialoside polymer as a candidate for a neuraminidase inhibitor showed that the glycopolymer has potent inhibitory activity against the neuraminidases. [Copyright &y& Elsevier]

Abstract: Carbosilane dendrimers periphery-functionalized with lactotriaose (GlcNAcβ1–3Galβ1–4Glc) with valencies of three, four, six, and twelve were prepared for use in a lectin-binding assay. A lactotriaose derivative was prepared from d-glucosamine and d-lactose derivatives. The N-Troc-protected glucosamine glycosyl donor and 3′-O-unprotected lactose glycosyl acceptor were condensed in the presence of silver trifluoromethanesulfonate and methylsulfenyl bromide to provide corresponding trisaccharide with new β-1-3 linkages in 92% yield. The protection group of the trisaccharide was transformed into an acetyl group. The 4-pentenyl glycoside was prepared from the acetate via glycosyl bromide. The alkene was converted into acetyl sulfide by addition of thioacetic acid under radical conditions. The lactotriaose unit was linked with carbosilane dendrimers to afford acetyl-protected glycodendrimers. De-O-acetylation of the dendrimers was carried out in the presence of sodium methoxide and then aq NaOH to give the desired lactotriaose clusters using a carbosilane dendrimer backbone. Their biological activities toward WGA were evaluated by fluorescence methods. The binding constants of free lactotriaose and trivalent, tetravalent, hexavalent, and dodecavalent glycodendrimers to WGA were determined to be 1.1×103, 4.4×104, 5.1×104, 2.8×106, and 1.3×106 M−1, respectively. The hexavalent glycodendrimer showed a 2500-fold larger binding effect than that of free lactotriaose. [Copyright &y& Elsevier]

Abstract: 4-Pentenyl (2,3,4,6-tetra-O-acetyl-β-d-galactopyranosyl)-(1→4)-(3,6-di-O-acetyl-2-deoxy-2-phthalimido-β-d-glucopyranosyl)-(1→3)-(2,6-di-O-benzoyl-β-d-galactopyranosyl)-(1→4)-2,3,6-tri-O-benzoyl-β-d-glucopyranoside (4) was synthesized by regioselective glycosylation of 4-pentenyl (2,6,-di-O-benzoyl-β-d-galactopyranosyl)-(1→4)-2,3,6-tri-O-benzoyl-β-d-glucopyranoside and (2,3,4,6-tetra-O-acetyl-β-d-galactopyranosyl)-(1→4)-3,6-di-O-acetyl-2-deoxy-2-phthalimido-β-d-glucopyranosyl chloride. By conversion of the protecting groups followed by thioacetylation, 4 was transformed into the corresponding lacto-N-neotetraose derivative, 5-(acetylthio)pentenyl (2,3,4,6-tetra-O-acetyl-β-d-galactopyranosyl)-(1→4)-O-(3,6-di-O-acetyl-2-acetamido-2-deoxy-β-d-glucopyranosyl)-(1→3)-(2,4,6-di-O-acetyl-β-d-galactopyranosyl)-(1→4)-2,3,6-tri-O-acetyl-β-d-glucopyranoside (6). The lacto-N-neotetraose derivative 6 was introduced into carbosilane dendrimer cores of three shapes, and three kinds of new carbosilane dendrimers peripherally functionalized by lacto-N-neotetraose were obtained. [Copyright &y& Elsevier]

Convenient access of a glycopolymer having N -acetyl- D -glucosamine (GlcNAc) residues was accomplished by means of radical polymerization of a simple GlcNAc monomer with or without acrylamide. Biological evaluations of glycopolymers displaying various carbohydrate contents were carried out by using fluorometric assays, and the results suggested that ratio of the sugar unit: acrylamide had a significant relation to affinities between carbohydrate—protein interactions. In this study, the unit ratio of 1:11.5 of carbohydrate: acrylamide was found to be the best ratio. Image 1 • A glycomonomer having GlcNAc was conventionally synthesized. • A series of glycopolymers having various densities of GlcNAc moieties was prepared. • Biological evaluations of glycopolymers—lectin were performed. • The contents of carbohydrate moieties—activity relationship of glycopolymers—lectin was investigated. [ABSTRACT FROM AUTHOR]

• Glycopolymer having GlcNAc moieties was conventionally prepared. • Efficient synthesis of glycopolymers having GlcNAc moieties and fluorophores was accomplished. • Biological evaluations of the glycopolymers—lectin were performed. • Intermolecular FRET was clearly observed using fluorogenic glycopolymers. A convenient assembly of fluorogenic glycopolymers having various polymer compositions was accomplished from the corresponding glycomonomer and dansyl monomer by means of radical polymerization, and the water-soluble glycopolymers gave typical fluorescence spectroscopic profiles due to the dansyl moieties on the glycopolymer in aqueous media. Biological evaluation of the polymer against wheat germ agglutinin (WGA) was accomplished on the basis of fluorescence changes due to tryptophan residues on WGA, and the affinities between the glycopolymers and WGA were estimated to be 4.7 × 105 to 9.3 × 105 M−1. In order to apply the fluorogenic glycopolymers for further biological measurements, efficient resonance energy transfer from tryptophan moieties on WGA to dansyl moieties on the fluorogenic glycopolymers was examined. FRET profiles of both fluorophores were similar compared to the binding profiles on the basis of fluorescence changes of tryptophan residues. This approach is applicable for the determination of an affinity constant between a carbohydrate and a lectin in which no fluorophore exists near the binding site. [ABSTRACT FROM AUTHOR]

• A prodrug-type neuraminidase substrate involving 4-nitroaniline was synthesized. • Neuraminidase activates the substrate to generate chromogenic 4-nitroaniline. • Neuraminidase is inactivated in the process of releasing 4-nitroaniline. Artificial substrates for probing neuraminidase activity are powerful tools for studying the physiological and pathological roles of neuraminidases. Most of the substrates are α- O -linked sialosides involving hydroxyl-containing reporters for visualization, and neuraminidase-catalyzed cleavage of the sialic acid residues directly activates the reporters. However, the use of amine-containing reporters has been avoided because α- N -linked sialosides are marginal substrates for neuraminidases. To expand the applicability of reporters to amine-containing compounds, we have focused on prodrug design. Herein we describe the synthesis and enzymatic study of a model substrate involving 4-nitroaniline as an amine-containing chromogenic reporter. The substrate can respond to neuraminidase from Clostridium perfringens. Neuraminidase-mediated hydrolysis of the sialic acid moiety of the substrate initiates self-immolative elimination of the linker moiety, leading the liberation of yellow-colored reporter 4-nitroaniline. The elimination process involves generation of quinone methide intermediate, which causes to neutralize neuraminidase. The substrate, thus, works as not only a chromogenic substrate but also a suicide inactivator. [ABSTRACT FROM AUTHOR]

[Display omitted] • Polyacrylamides functionalized with amino acids were synthesized. • Some polymers showed cleavage activity against bovine and human serum albumins. • The cleavage profiles of the active polymers differed from those of α-chymotrypsin. • At 50 °C, α-chymotrypsin lost activity, but the polymer showed better protein cleavage. Polyacrylamides with various compositions of serine, aspartic acid, and histidine, which are the amino acids involved in the catalytic triad of natural serine protease chymotrypsin, were synthesized and their protein cleavage activity was investigated. SDS-PAGE analysis showed that some of the synthesized ternary copolymers showed cleavage activity against bovine and human serum albumins. Polyacrylamides incorporating a single type of amino acid were also able to cleave the protein substrates. These homopolymers exhibited unique cleavage profiles and pH and temperature sensitivities that differed from those of α-chymotrypsin. The results indicate the potential of polymers functionalized with amino acids as proteolytic artificial enzymes. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Sugar-amino acid hybrid monomers were prepared. • Assembly of the monomers was accomplished by radical polymerization. • Affinities of the polymers against Stxs were examined. • High cluster-type polymers showed higher affinity than low cluster-type polymers. Abstract Synthetic assembly of sugar moieties and amino acids in order to create "sugar-amino acid hybrid polymers" was accomplished by means of simple radical polymerization of carbohydrate monomers having an amino acid-modified polymerizable aglycon. Amines derived from globotriaoside and lactoside as glycoepitopes were condensed with known carbobenzyloxy derivatives, including Z-Gly, Z- l -Ala and Z-β-Ala, which had appropriate spacer ability and a chiral center to afford fully protected sugar-amino acid hybrid compounds in good yields. After deprotection followed by acryloylation, the water-soluble glycomonomers were polymerized with or without acrylamide in the presence of a radical initiator in water to give corresponding copolymers and homopolymers, which were shown by SEC analysis to have high molecular weights. Evaluation of the biological activities of the glycopolymers against Shiga toxins (Stxs) was carried out, and the results suggested that glycopolymers having highly clustered globotriaosyl residues had high affinity against Stx2 (K D = 2.7∼4.0 µM) even though other glycopolymers did not show any affinity or showed very weak binding affinity. When Stx1 was used for the same assay, all of the glycopolymers having globotriaosyl residues showed high affinity (K D = 0.30∼1.74 µM). Interestingly, couple of glycopolymers having lactosyl moieties had weaker binding affinity against Stx1. In addition, when cytotoxicity assays were carried out for both Stxs, glycopolymers having highly clustered globotriaosyl residues showed higher affinity than that of the copolymers, and only highly clustered-type glycopolymers displayed neutralization potency against Stx2. [ABSTRACT FROM AUTHOR]

[Display omitted] • NMR analyses of CNN and its derivatives were performed. • Verification of standard acetylation for CNN was performed. • Complete protection of CNN was efficiently performed. Verification of acetylation by means of pyr—Ac 2 O for a cyclic α-nigerosyl-(1 → 6)-nigerose (CNN) was performed and the corresponding mono alcohol was obtained. Structural elucidation of CNN derivatives was carried out on the basis of NMR spectroscopic analyses. Complete protection by means of esterification and methylation was efficiently accomplished to yield fully protected compounds, and the results of NMR spectroscopic analyses of the products supported the perfect progress of the reaction. [ABSTRACT FROM AUTHOR]

The carbohydrate moieties of arabinogalactan-proteins (AGPs) have β-(1→3)-galactan backbones to which side chains of (1→6)-linked β-Gal residues are attached through O -6. Some of these side chains are further substituted with other sugars. We investigated the structure of l -Fuc-containing oligosaccharides released from the carbohydrate moieties of a radish leaf AGP by digestion with α- l -arabinofuranosidase, followed by exo-β-(1→3)-galactanase. We detected a series of neutral β-(1→6)-galactooligosaccharides branching variously at O -3 of the Gal residues, together with corresponding acidic derivatives terminating in 4- O -methyl-GlcA (4-Me-GlcA) or GlcA at the non-reducing terminals. In neutral oligosaccharides with degree of polymerization (dp) mainly higher than 10, l -Fuc groups were attached through l -Ara residues as the sequence, α- l -Fuc p -(1→2)-α- l -Ara f -(1→. This sequence was verified by isolation of the pentasaccharide α- l -Fuc-(1→2)-α- l -Ara f -(1→3)-β-Gal-(1→6)-β-Gal-(1→6)-Gal upon digestion of the higher oligosaccharides with endo-β-(1→6)-galactanase. By contrast, in lower polymerized (predominantly dp 4) acidic oligosaccharides, l -Fuc groups were attached directly at the non-reducing terminals through α-(1→2)-linkages, resulting in the release of the tetrasaccharides, α- l -Fuc p -(1→2)-β-GlcA-(1→6)-β-Gal-(1→6)-Gal and α- l -Fuc p -(1→2)-β-4-Me-GlcA-(1→6)-β-Gal-(1→6)-Gal. In long acidic oligosaccharides with dp mainly higher than 13, l -Fuc groups localized on branches were attached to the uronic acids directly and/or l -Ara residues as in the neutral oligosaccharides. [ABSTRACT FROM AUTHOR]

[Display omitted] • Glycopolymer having sialyl α 2–3 lactose moieties was prepared. • Sugar densities on the glycopolymers were controlled. • Biological evaluations of glycopolymers—MuV were tested. • Glycopolymer having low sugar-density showed the highest affinity. A water-soluble glycomonomer having a sialyl α2 → 3 lactose (SLac) moiety was prepared from a known imidate derivative of the SLac and an acrylamide alcohol by means of Schmidt's protocol followed by transesterification. Polymerization of the monomer proceeded in water as the solvent in the presence of ammonium persulfate (APS)—tetramethylethylenediamine (TEMED). Since acryl amide (AAm) was used as a regulator for the arrangement of sugar density, three kinds of glycopolymers having different sugar densities were obtained. Infection inhibition assays of mumps virus (MuV) for Vero cells using the glycopolymers were performed, and the results showed that a glycopolymer having a low sugar density has the highest inhibitory potency. In comparison to sialyl Lewis X (SLeX) as the strongest inhibitor in a previous study, SLac polymer with the low sugar density showed ten-times stronger inhibitory potency than that of SLex. This finding suggested that multivalent conversion of the monomeric SLac with appropriate spatial arrangement are able to effectively inhibit the interaction between the attachment glycoprotein of MuV and glycan receptors on Vero cells. [ABSTRACT FROM AUTHOR]

Abstract: The LeX glycoside having an N3 moiety at the ω position, which was prepared by using living cells from GlcNAcO(CH2)12N3 as a starting material, was efficiently introduced into a trivalent-type carbosilane core scaffold by means of Huisgen cycloaddition reaction to yield the corresponding glycocluster having three LeX moieties at each end. Structural elucidation of the product was performed by a combination of NMR and mass spectroscopic analyses, and the results of the analyses supported the structure of the glycocluster. Evaluation of the glycocluster was carried out using Lotus lectin as a model lectin. Environmental changes of tryptophan residues located at or near the binding sites of Lotus lectin caused fluorescence intensity change. [Copyright &y& Elsevier]

Abstract: A new compound having a 2,3,4,5-tetraphenylsilole derivative on the center silicon of Dumbbell(1)6Gb3; Silole-Dumbbell(1)6Gb3 (1) was previously reported. It was found that 1 exhibited strongly increased fluorescence both in water and in a 96% acetone/water mixed solvent. The physical behavior of 1 in water and in the 96% acetone/water mixed solvent was investigated, and analyses including fluorescence quantum yields, dynamic-light-scattering (DLS), atomic-force-microscopy (AFM), and fluorescence microscopy were carried out. It was clarified that 1 dynamically formed different types of aggregates in water and in higher acetone concentrations to yield high aggregation-induced emission (AIE) effects due to the formation of micelle-like particles in water and inversion-type micelles in the acetone/water mixed solvent, respectively. [Copyright &y& Elsevier]

Abstract: A glycocluster peripherally functionalized with a lactose (Lac: Galβ1→4Glcβ1–) derivative possessing a silole moiety as a luminophore was synthesized. The photoluminescence spectrum of the glycocluster showed extremely strong emission at 474nm and the absolute quantum yield was estimated to be 92% in distilled water. The emission intensity was decreased by increasing the amount of peanut agglutinin (PNA), a lactose-binding lectin, and plots of the relative fluorescence intensity revealed a decline of 95% in emission intensity. Fluorescence quenching of the glycocluster upon mixing with PNA could be easily observed by the naked eye under UV irradiation, whereas no distinct change in fluorescence properties of the glycocluster was observed when wheat germ agglutinin (WGA) was employed. [Copyright &y& Elsevier]

Abstract: Synthetic construction of a Lex determinant was accomplished via slightly modified Gabriel amine synthesis from three building blocks. Further transformations followed by polymerization of the Lex derivative gave a glycopolymer having trisaccharidic units as pendant-type epitopes. [Copyright &y& Elsevier]

Abstract: A novel O-protected lactose (Galβ1→4Glcβ1–) derivative bearing trimethoxysilyl group at the aglycon was developed as a silane coupling agent. Reaction of the coupling agent with tris(2,2′-bipyridine)ruthenium (II) dichloride (Rubpy) doped silica particle gave a Rubpy-doped silica particle peripherally functionalized with O-protected lactose derivative. De-O-protection of the particle with aqueous ammonia provided lactose-coating Rubpy-doped silica particles, combining luminophor encapsulated in silica matrix and carbohydrate having lectin-recognition ability. Specific adhesion of fluorescein isothiocyanate-labeled peanut agglutinin (FITC-PNA) to the lactose-coating Rubpy-doped silica particles was confirmed by fluorescence microscopic analysis. [Copyright &y& Elsevier]

Abstract: The carbohydrate moieties of arabinogalactan-proteins (AGPs), which are mainly composed of Gal, l-Ara, GlcA, and 4-Me-GlcA residues, are essential for the physiological functions of these proteoglycans in higher plants. For this study, we have identified two genes encoding family 79 β-glucuronidases, designated AnGlcAase and NcGlcAase, in Aspergillus niger and Neurospora crassa, respectively, based on the amino acid sequence of a native β-glucuronidase purified from a commercial pectolytic enzyme preparation from A. niger. Although the deduced protein sequences of AnGlcAase and NcGlcAase were highly similar, the recombinant enzymes expressed in Pichia pastoris exhibited distinct substrate specificity toward 4-Me-GlcA residues of AGPs: recombinant AnGlcAase (rAnGlcAase) substantially liberated both GlcA and 4-Me-GlcA residues from radish AGPs, whereas recombinant NcGlcAase (rNcGlcAase) activity on the 4-Me-GlcA residues of AGPs was very low. Maximum activity of rAnGlcAase hydrolyzing PNP β-GlcA occurred at pH 3.0–4.0, whereas the maximum rNcGlcAase activity was at pH 6.0. The apparent K m values of rAnGlcAase were 30.4μM for PNP β-GlcA and 422μM for β-GlcA-(1→6)-Gal, and those of rNcGlcAase were 38.3μM and 378μM, respectively. Similar to the native enzyme, rAnGlcAase was able to catalyze the transglycosylation of GlcA residues from PNP β-GlcA to various monosaccharide acceptors such as Glc, Gal, and Xyl. We propose that both AnGlcAase and NcGlcAase are instances of a novel type of β-glucuronidase with the capacity to hydrolyze β-GlcA and 4-Me-β-GlcA residues of AGPs, although they differ significantly in their preferences. [Copyright &y& Elsevier]

Monomeric sugars generated during the metabolism of polysaccharides, glycoproteins, and glycolipids are imported to the cytoplasm and converted to respective nucleotide sugars via monosaccharide 1-phosphates, to be reutilized as activated sugars. Because L-fucose (L-Fuc) is activated mainly in the form of GDP derivatives in seed plants, the salvage reactions for L-Fuc are expected to be independent from those for GIc, Gal, L-arabinose, and glucuronic acid, which are activated as UDP-sugars. For this study we have identified, in the genomic data base of Arabidopsis, the gene (designated AtFKGP) of a bifunctional enzyme with similarity to both L-fucokinase and GDP-L-Fuc pyrophosphorylase. Recombinant AtFKGP (rAt-FKGP) expressed in Escherichia coli showed both L-fucokinase and GDP-L-Fuc pyrophosphorylase activities, generating GDP- L-Fuc from L-Fuc, ATP, and GTP as the starting substrates. Point mutations in rAtFKGPs at either Gly133 or Gly830 caused loss of GDP-L-Fuc pyrophosphorylase and L-fucokinase activity, respectively. The apparent Km, values of L-fucokinase activity of rAtFKGP for L-Fuc and ATP were 1.0 and 0.45 mM, respectively, and those of GDP-L-Fuc pyrophosphorylase activity for L-Fuc 1-phosphate and GTP were 0.052 and 0.17 mM, respectively. The expression of AtFKGP was detected in most cell types of Arabidopsis, indicating that salvage reactions for free L-Fuc catalyzed by AtFKGP occur ubiquitously in Arabidopsis. Loss-of-function mutants with tDNA insertion in AtFKGP exhibited higher accumulation of free L-Fuc in the soluble fraction than the wild-type plant. These results indicate that AtFKGP is a bifunctional enzyme with L-fucokinase and GDP-L-Fuc pyrophosphorylase activities, which salvages free L-Fuc in Arabidopsis. [ABSTRACT FROM AUTHOR]

Abstract: A novel glycocluster periphery functionalized by globotriaose (Galα1–4Galβ1–4Glcβ1–) possessing a silole moiety as a luminophor was synthesized. The photoluminescence spectrum of the glycocluster in pure water showed extremely strong emission at 475nm. Analogous intense emission of the silole dendrimer was also observed in a lower water fraction of water/acetone mixture. The water fraction of the silole dendrimer solution strongly affected the emission intensity; however, these luminescences were constantly detected at around 475nm. [Copyright &y& Elsevier]

Convenient and useful construction of a trisaccharide moiety of globotriaosyl ceramide was performed by means of modified Ogawa’s protocol. In order to evaluate an efficiency of new class of glycopolymers, further chemical transformations of the trisaccharide were accomplished to afford a globotriaosyl carbohydrate monomer, and homopolymerization of the monomer by a general radical polymerization protocol gave a high-density glycopolymer in 84.3% yield after usual work-up procedures. In addition to the homopolymer, a copolymer composed of carbohydrate units and acrylamide units was also synthesized by the radical polymerization in 96.7% yield. [Copyright &y& Elsevier]

Abstract: Carbosilane dendrimers bearing galabiose (Galα1–4Gal) with three, four, and six galabiose units at the periphery of the dendrimers were synthesized for use as artificial inhibitors against Shiga toxins (Stxs) produced by Escherichia coli O157:H7. The galabiose unit, prepared from penta-O-acetyl-β-d-galactopyranose, was linked with carbosilane dendrimers of three shapes to afford acetyl-protected glycodendrimers in good yields. De-O-acetylation of the clusters was carried out in the presence of NaOMe and then aq NaOH to give the desired three shapes of galabiose-coated carbosilane dendrimers. Their biological activities toward Stxs were evaluated by kinetic analysis, binding assays, and cytotoxic assays. [Copyright &y& Elsevier]

As an artificial ligand, a glycoconjugate polymer carrying carbohydrate moiety of lactosyl ceramide or globotriaosyl ceramide (Gb3) was synthesized. Gb3 is known as the receptor of Shiga toxin-producing Escherichia coli O157: H7. The preparation of the glycoconjugate polymer initially involves the construction of the carbohydrate moiety of Gb3 derivative which has n-pentenyl group as polymerizable group. In addition, the n-pentenyl group of the Gb3 derivative was modified and different polymerizable groups such as acrylamide group were introduced at ω-position of the aglycon. Radical polymerization of the synthesized glycosyl monomers with or without acrylamide proceeded smoothly in water using ammonium persulfate and N, N, N′, N′-tetramethylethylenediamine as usual initiator system and gave water-soluble glycoconjugate polymers having various polymer compositions. These polymers have the potential to neutralize Shiga toxin by reason of cluster effect and multivalency. [Copyright &y& Elsevier]

A novel anomeric β-thioacetate of an N-acetyllactosamine derivative was efficiently synthesized in high yield from the known 2-azido glycosyl chloride using thioacetic acid as a convenient reagent. The synthesis involved not only an SN2 replacement of the chloride by a carbothiolate anion but also a reductive acetamidation of the azide group. Applications of the thioacetate for glycosidation were demonstrated to provide both O- and S-glycosides in high yields. Furthermore, both intermediates gave a new class of glycoclusters that included thioglycosidic linkages. [Copyright &y& Elsevier]