Highly Modified Cellulose Nanocrystals and Formation of Epoxy-Nanocrystalline Cellulose (CNC) Nanocomposites.

This work presents an environmentally friendly, iodine-catalyzed chemical modification method to generate highly hydrophobic, optically active nanocrystalline cellulose (CNC). The high degree of ester substitution (DS = 2.18), hydrophobicity, crystalline behavior, and optical activity of the generated acetylated CNC (Ac-CNC) were quantified by TEM, FTIR, solid 13C NMR, contact angle, XRD, and POM analyses. Ac-CNC possesses substantial enhancement in thermal stability (16.8%) and forms thin films with an interlayer distance of 50-150 nm, presenting cavities suitable for entrapping nano- and microparticles. Generated Ac-CNC proved to be an effective reinforcing agent in hydrophobic polymer matrices for fabricating high performance nanocomposites. When integrated at a very low weight percentage (0.5%) in an epoxy matrix, Ac-CNC provided for a 73% increase in tensile strength and a 98% increase in modulus, demonstrating its remarkable reinforcing potential and effective stress transfer behavior. The method of modification and the unique properties of the modified CNC (hydrophobicity, crystallinity, reinforcing ability, and optical activity) render them a novel bionanomaterial for a range of multipurpose applications.

A general route to xyloglucan-peptide conjugates for the activation of cellulose surfaces.

Cellulose is an attractive supporting matrix for diverse biotechnological applications, including chromatography, diagnostics, and tissue replacement/scaffolding, due to its renewable resource status, low cost, and low non-specific interaction with biomolecules. In an effort to expand the biofunctionality of cellulose materials, we present here a versatile method for the synthesis of xyloglucan-peptide conjugates that harness the strong xyloglucan-cellulose binding interaction for gentle surface modification. Xylogluco-oligosaccharide aminoalditols (XGO-NH(2)) were coupled to both linear and cyclic peptides, which contained the endothelial cell epitope Arg-Gly-Asp, in a facile two-step approach employing diethyl squarate cross-linking. Subsequent xyloglucan endo-transglycosylase-mediated coupling of the resulting XGO-GRGDS (Gly-Arg-Gly-Asp-Ser) and XGO-c[RGDfK]-PEG-PEG (cyclo[Arg-Gly-Asp-(D-Phe)-Lys]-PEG-PEG; where PEG is 8-amino-3,6-dioxaoctanoic acid) conjugates into high molecular mass xyloglucan yielded xyloglucan-RGD peptide conjugates suitable for cellulose surface activation. Notably, use of XGO-squaramate as a readily accessible, versatile intermediate overcomes previous limitations of solid-phase synthetic approaches to XGO-peptide conjugates, and furthermore allows the method to be generalized to a wide variety of polypeptides and proteins, as well as diverse primary amino compounds.

Intervening with urinary tract infections using anti-adhesives based on the crystal structure of the FimH-oligomannose-3 complex.

BACKGROUND: Escherichia coli strains adhere to the normally sterile human uroepithelium using type 1 pili, that are long, hairy surface organelles exposing a mannose-binding FimH adhesin at the tip. A small percentage of adhered bacteria can successfully invade bladder cells, presumably via pathways mediated by the high-mannosylated uroplakin-Ia and alpha3beta1 integrins found throughout the uroepithelium. Invaded bacteria replicate and mature into dense, biofilm-like inclusions in preparation of fluxing and of infection of neighbouring cells, being the major cause of the troublesome recurrent urinary tract infections. METHODOLOGY/PRINCIPAL FINDINGS: We demonstrate that alpha-D-mannose based inhibitors of FimH not only block bacterial adhesion on uroepithelial cells but also antagonize invasion and biofilm formation. Heptyl alpha-D-mannose prevents binding of type 1-piliated E. coli to the human bladder cell line 5637 and reduces both adhesion and invasion of the UTI89 cystitis isolate instilled in mouse bladder via catheterization. Heptyl alpha-D-mannose also specifically inhibited biofilm formation at micromolar concentrations. The structural basis of the great inhibitory potential of alkyl and aryl alpha-D-mannosides was elucidated in the crystal structure of the FimH receptor-binding domain in complex with oligomannose-3. FimH interacts with Man alpha1,3Man beta1,4GlcNAc beta1,4GlcNAc in an extended binding site. The interactions along the alpha1,3 glycosidic bond and the first beta1,4 linkage to the chitobiose unit are conserved with those of FimH with butyl alpha-D-mannose. The strong stacking of the central mannose with the aromatic ring of Tyr48 is congruent with the high affinity found for synthetic inhibitors in which this mannose is substituted for by an aromatic group. CONCLUSIONS/SIGNIFICANCE: The potential of ligand-based design of antagonists of urinary tract infections is ruled by the structural mimicry of natural epitopes and extends into blocking of bacterial invasion, intracellular growth and capacity to fluxing and of recurrence of the infection.

Synthesis of fused bicyclic thioglycosides of N-acylated glucosamine as analogues of mycothiol.

The synthesis of a fused bicyclic thioglycoside analogue of mycothiol, (3R)-3-acetylamino-4-one-6,7-dihydro-(1',2'-dideoxy-beta-D-glucopyranoso)[2',1'-f]-1,5-thiazepane (5), is reported. Treatment of phthalimido-protected peracetylated glucosamine with N-acetyl-cysteine and boron trifluoride-etherate gave the beta-linked thioglycoside, which was deprotected and cyclized, using HOBt and EDCl to form the lactam and giving the target structure. This mycothiol mimic and its tri-O-acetate will be investigated as potential inhibitors of enzymes involved in the biosynthesis of mycothiol. The protected derivative also has the potential to be an alpha-selective N-cysteinyl glucosamine donor; however, initial glycosylation attempts failed due to the apparent stability of the fused bicyclic system.

Synthesis of stable C-phosphonate analogues of Neisseria meningitidis group A capsular polysaccharide structures using modified Mitsunobu reaction conditions.

Examples of synthetic C-phosphonate analogues of microbial polysaccharide structures containing inter-residue phosphodiester linkages are most rare. The successful construction of such analogues of the Neisseria meningitidis Group A capsular polysaccharide is described. Using a modified Mitsunobu reaction (tris(4-chlorophenyl)phosphine, DIAD, excess of Et3N) between an anomeric C-phosphonate monoester and a 6-OH ManNAc acceptor a high yield (88%) of a dimer was obtained. Transformation of the dimer into a new 6-OH acceptor through deacetylation and further reaction with the elongating C-phosphonate monomer employing the same conditions afforded the trimer in 92% yield. Iteration of the procedure then afforded the tetramer with a coupling yield of 85%. The di-, tri- and tetramer were deprotected to give target structures ready for conjugation to a carrier protein and subsequent immunological evaluation.

Synthesis of structures corresponding to the capsular polysaccharide of Neisseria meningitidis group A.

Four differently substituted trimers of the CPS repeating unit have been synthesised in order to investigate the dependence on oligosaccharide size, acetylation and mode of phosphorylation of glycoconjugate vaccines against Neisseria meningitidis group A. A spacer-containing starting monomer, a H-phosphonate elongating monomer and a 6-O-phosphorylated H-phosphonate cap monomer have been synthesised and coupled together to afford, after deprotection, the target trimer structures differing in their acetylation and phosphorylation substitution pattern.

Improved synthesis of 1,3,4,6-tetra-O-acetyl-2-azido-2-deoxy-alpha-D-mannopyranose.

By improved (anhydrous) work-up conditions of a triflate displacement reaction, the yield in the preparation of the versatile synthetic intermediate 1,3,4,6-tetra-O-acetyl-2-azido-2-deoxy-alpha-d-mannopyranose has been significantly enhanced. This important precursor is now available in three efficient steps from d-glucose.

Banana lectin is unique in its recognition of the reducing unit of 3-O-beta-glucosyl/mannosyl disaccharides: a calorimetric study.

The binding of banana lectin (BanLec) to laminaribiose (Glcbeta1,3Glc) and a series of novel synthetic analogues was measured by titration calorimetry to assess the contribution of the hydroxyl groups of the reducing glycosyl moiety and its 3-O-beta-substituent to binding. Key areas of interaction involved the 1, 2, and 6 positions of the reducing-terminal hexose unit. The alpha-anomeric configuration of the reducing hexose was strongly favored over the beta-anomer. The 2-hydroxyl in the axial position (mannose) also enhanced binding, whereas the 6-hydroxymethyl group was essential, because xylopyranose in the reducing position was inactive. The 3-O-beta-glucosyl unit of methyl alpha-laminaribioside could be replaced by any of its monodeoxy derivatives. However, the 4'-deoxy derivative or axial hydroxy (galactosyl) substitution was somewhat detrimental to binding. 3-O-substitution with the (S)tetrahydropyranyl ring or a benzyl group had similar effect as 4'-deoxyglucosyl substitution. Surprisingly, p-nitrobenzyl or beta-xylosyl 3-O-substitution greatly enhanced binding of the reducing glucosyl or mannosyl derivative. Chemical syntheses of a number of novel disaccharides and analogues prepared for this study are described.

Receptor binding studies disclose a novel class of high-affinity inhibitors of the Escherichia coli FimH adhesin.

Mannose-binding type 1 pili are important virulence factors for the establishment of Escherichia coli urinary tract infections (UTIs). These infections are initiated by adhesion of uropathogenic E. coli to uroplakin receptors in the uroepithelium via the FimH adhesin located at the tips of type 1 pili. Blocking of bacterial adhesion is able to prevent infection. Here, we provide for the first time binding data of the molecular events underlying type 1 fimbrial adherence, by crystallographic analyses of the FimH receptor binding domains from a uropathogenic and a K-12 strain, and affinity measurements with mannose, common mono- and disaccharides, and a series of alkyl and aryl mannosides. Our results illustrate that the lectin domain of the FimH adhesin is a stable and functional entity and that an exogenous butyl alpha-D-mannoside, bound in the crystal structures, exhibits a significantly better affinity for FimH (Kd = 0.15 microM) than mannose (Kd = 2.3 microM). Exploration of the binding affinities of alpha- d-mannosides with longer alkyl tails revealed affinities up to 5 nM. Aryl mannosides and fructose can also bind with high affinities to the FimH lectin domain, with a 100-fold improvement and 15-fold reduction in affinity, respectively, compared with mannose. Taken together, these relative FimH affinities correlate exceptionally well with the relative concentrations of the same glycans needed for the inhibition of adherence of type 1 piliated E. coli. We foresee that our findings will spark new ideas and initiatives for the development of UTI vaccines and anti-adhesive drugs to prevent anticipated and recurrent UTIs.