Stereoselective synthesis of C-glycosides from carboxylic acids: the tandem Tebbe-Claisen approach

A variety of beta- or alpha-C-glycosides may be readily accessed in an entirely stereoselective fashion from esters derived from the reaction of carboxylic acids and 3-hydroxy glycals, by way of a tandem reaction sequence of Tebbe methylenation and Claisen rearrangement. Though of wide scope, for example allowing the synthesis of 1-6 linked C-disaccharides, the methodology does not currently allow the synthesis of C-glycosyl alpha-amino acids.     

Allyl Protecting Group Mediated Intramolecular Aglycon Delivery (IAD) of Glycosyl Fluorides

Summary.  Stereospecific 1,2-cis-glycosylation of 2-O-allyl protected glucosyl and mannosyl fluorides can be achieved via a sequence of allyl isomerization, N-iodosuccinimide mediated tethering, and intramolecular aglycon delivery (IAD). Fluoride is advantageous as an anomeric leaving group since extended reaction times can be employed to tether hindered aglycon alcohols without competitive anomeric activation. Tin(II) chloride mediated intramolecular glycosylation furnishes the desired α-glucosides and β-mannosides in an entirely stereoselective manner. Received August 24, 2001. Accepted November 6, 2001     

Chemoenzymatic Synthesis of a Phosphorylated Glycoprotein

The majority of lysosomal enzymes are targeted to the lysosome by post‐translational tagging with N‐glycans terminating in mannose‐6‐phosphate (M6P) residues. Some current enzyme replacement therapies (ERTs) for lysosomal storage disorders are limited in their efficacy by the extent to which the recombinant enzymes bear the M6P‐terminated glycans required for effective trafficking. Chemical synthesis was combined with endo‐β‐N‐acetylglucosaminidase (ENGase) catalysis to allow the convergent synthesis of glycosyl amino acids bearing M6P residues. This approach can be extended to the remodeling of proteins, as exemplified by RNase. The powerful synergy of chemical synthesis and ENGase‐mediated biocatalysis enabled the first synthesis of a glycoprotein bearing M6P‐terminated N‐glycans in which the glycans are attached to the peptide backbone by entirely natural linkages.     

Characterization of High Voltage Cold Atmospheric Plasma Generation in Sealed Packages as a Function of Container Material and Fill Gas

While numerous experiments have demonstrated the efficacy of high voltage cold atmospheric pressure plasmas (HVCAPs) in sealed packages for microbial inactivation, the influence of the package on the emitted species measured during HVCAP discharge is poorly understood. This study elucidates the impact of the package on plasma generation in sealed packages for four separate gases (ambient air, commercial grade compressed air, a helium/air mixture, and nitrogen) placed in commercially available transparent plastic containers and bags representative of the materials used in the food industry. The container and bag individually reduced emission signal intensity by an average of 63 and 45%, respectively, across the measured wavelengths of 200–1100 nm, demonstrating that they acted as broadband absorbers. Neither the container nor bag caused additional emission lines to appear, indicating no significant effect on the types of species generated. Considering the minimum applied voltage necessary to induce a discharge, the power dissipated by the nitrogen and ambient air plasma generated at 72 ± 3.7 kV RMS were comparable to the compressed dry air discharge generated at 80 ± 3.7 kV RMS. The helium discharge at 37 ± 3.7 kV RMS absorbed approximately 92% more power than these gases. Rotational temperatures ranged from 285 K for helium to 479 K for compressed air. These results indicate that the package impacts the intensity distribution but not the presence of the most dominant peaks, although further studies are required to elucidate the impact on less intense peaks.     

Convergent chemo-enzymatic synthesis of mannosylated glycopeptides; targeting of putative vaccine candidates to antigen presenting cells

The combination of solid phase peptide synthesis and endo-β-N-acetylglucosaminidase (ENGase) catalysed glycosylation is a powerful convergent synthetic method allowing access to glycopeptides bearing full-length N-glycan structures. Mannose-terminated N-glycan oligosaccharides, produced by either total or semi-synthesis, were converted into oxazoline donor substrates. A peptide from the human cytomegalovirus (CMV) tegument protein pp65 that incorporates a well-characterised T cell epitope, containing N-acetylglucosamine at specific Asn residues, was accessed by solid phase peptide synthesis, and used as an acceptor substrate. High-yielding enzymatic glycosylation afforded glycopeptides bearing defined homogeneous high-mannose N-glycan structures. These high-mannose containing glycopeptides were tested for enhanced targeting to human antigen presenting cells (APCs), putatively mediated via the mannose receptor, and for processing by the APCs for presentation to human CD8+ T cells specific for a 9-mer epitope within the peptide. Binding assays showed increased binding of glycopeptides to APCs compared to the non-glycosylated control. Glycopeptides bearing high-mannose N-glycan structures at a single site outside the T cell epitope were processed and presented by the APCs to allow activation of a T cell clone. However, the addition of a second glycan within the T cell epitope resulted in ablation of T cell activation. We conclude that chemo-enzymatic synthesis of mannosylated glycopeptides enhances uptake by human APCs while preserving the immunogenicity of peptide epitopes within the glycopeptides, provided those epitopes are not themselves glycosylated.     

Experimental investigation of the breakdown of the Onsager-Casimir relations

We use magnetoconductance fluctuation measurements of phase-coherent semiconductor billiards to quantify the contributions to the nonlinear electric conductance that are asymmetric under reversal of magnetic field. We find that the average asymmetric contribution is linear in magnetic field (for magnetic flux much larger than 1 flux quantum) and that its magnitude depends on billiard geometry. In addition, we find an unexpected asymmetry in the power spectrum of the magnetoconductance with respect to reversal of magnetic field and bias voltage.     

The importance of including local correlation times in the calculation of inter-proton distances from NMR measurements: ignoring local correlation times leads to significant errors in the conformational analysis of the Glc alpha1-2Glc alpha linkage by NMR spectroscopy

The experimental determination of oligosaccharide conformations has traditionally used cross-linkage 1H-1H NOE/ROEs. As relatively few NOEs are observed, to provide sufficient conformational constraints this method relies on: accurate quantification of NOE intensities (positive constraints); analysis of absent NOEs (negative constraints); and hence calculation of inter-proton distances using the two-spin approximation. We have compared the results obtained by using 1H 2D NOESY, ROESY and T-ROESY experiments at 500 and 700 MHz to determine the conformation of the terminal Glc alpha1-2Glc alpha linkage in a dodecasaccharide and a related tetrasaccharide. For the tetrasaccharide, the NOESY and ROESY spectra produced the same qualitative pattern of linkage cross-peaks but the quantitative pattern, the relative peak intensities, was different. For the dodecasaccharide, the NOESY and ROESY spectra at 500 MHz produced a different qualitative pattern of linkage cross-peaks, with fewer peaks in the NOESY spectrum. At 700 MHz, the NOESY and ROESY spectra of the dodecasaccharide produced the same qualitative pattern of peaks, but again the relative peak intensities were different. These differences are due to very significant differences in the local correlation times for different proton pairs across this glycosidic linkage. The local correlation time for each proton pair was measured using the ratio of the NOESY and T-ROESY cross-relaxation rates, leaving the NOESY and ROESY as independent data sets for calculating the inter-proton distances. The inter-proton distances calculated including the effects of differences in local correlation times give much more consistent results.