Automatic Structure Determination of Regular Polysaccharides Based Solely on NMR Spectroscopy

The structural analysis of polysaccharides requires that the sugar components and their absolute configurations are determined. We here show that this can be performed based on NMR spectroscopy by utilizing butanolysis with (+)- and (−)-2-butanol that gives the corresponding 2-butyl glycosides with characteristic ¹H and ¹³C NMR chemical shifts. The subsequent computer-assisted structural determination by CASPER can then be based solely on NMR data in a fully automatic way as shown and implemented herein. The method is additionally advantageous in that reference data only have to be prepared once and from a user’s point of view only the unknown sample has to be derivatized for use in CASPER

Conformational Preferences of the O‑Antigen Polysaccharides of Escherichia coli O5ac and O5ab Using NMR Spectroscopy and Molecular Modeling

Escherichia coli serogroup O5 comprises two different subgroups (O5ab and O5ac), which are indiscernible from the point of view of standard immunological serotyping. The structural similarities between the O-antigen polysaccharides (PSs) of these two strains are remarkable, with the only difference being the glycosidic linkage connecting the biological tetrasaccharide repeating units. In the present study, a combination of NMR spectroscopy and molecular modeling methods were used to elucidate the conformational preferences of these two PSs. The NMR study was based on the analysis of intra- and inter-residue proton–proton distances using NOE build-up curves. Molecular models of the repeating units and their extension to polysaccharides were obtained, taking into account the conformational flexibility as assessed by the force field applied and a genetic algorithm. The agreements between experimentally measured and calculated distances could only be obtained by considering an averaging of several low energy conformations observed in the molecular models

Conformational Preferences of the O‑Antigen Polysaccharides of Escherichia coli O5ac and O5ab Using NMR Spectroscopy and Molecular Modeling

Escherichia coli serogroup O5 comprises two different subgroups (O5ab and O5ac), which are indiscernible from the point of view of standard immunological serotyping. The structural similarities between the O-antigen polysaccharides (PSs) of these two strains are remarkable, with the only difference being the glycosidic linkage connecting the biological tetrasaccharide repeating units. In the present study, a combination of NMR spectroscopy and molecular modeling methods were used to elucidate the conformational preferences of these two PSs. The NMR study was based on the analysis of intra- and inter-residue proton–proton distances using NOE build-up curves. Molecular models of the repeating units and their extension to polysaccharides were obtained, taking into account the conformational flexibility as assessed by the force field applied and a genetic algorithm. The agreements between experimentally measured and calculated distances could only be obtained by considering an averaging of several low energy conformations observed in the molecular models

Conformational Preferences of the O‑Antigen Polysaccharides of Escherichia coli O5ac and O5ab Using NMR Spectroscopy and Molecular Modeling

Escherichia coli serogroup O5 comprises two different subgroups (O5ab and O5ac), which are indiscernible from the point of view of standard immunological serotyping. The structural similarities between the O-antigen polysaccharides (PSs) of these two strains are remarkable, with the only difference being the glycosidic linkage connecting the biological tetrasaccharide repeating units. In the present study, a combination of NMR spectroscopy and molecular modeling methods were used to elucidate the conformational preferences of these two PSs. The NMR study was based on the analysis of intra- and inter-residue proton–proton distances using NOE build-up curves. Molecular models of the repeating units and their extension to polysaccharides were obtained, taking into account the conformational flexibility as assessed by the force field applied and a genetic algorithm. The agreements between experimentally measured and calculated distances could only be obtained by considering an averaging of several low energy conformations observed in the molecular models

Chemoenzymatic Synthesis of <i>O</i>-Mannosylpeptides in Solution and on Solid Phase

<i>O</i>-Mannosyl glycans are known to play an important role in regulating the function of α-dystroglycan (α-DG), as defective glycosylation is associated with various phenotypes of congenital muscular dystrophy. Despite the well-established biological significance of these glycans, questions regarding their precise molecular function remain unanswered. Further biological investigation will require synthetic methods for the generation of pure samples of homogeneous glycopeptides with diverse sequences. Here we describe the first total syntheses of glycopeptides containing the tetrasaccharide NeuNAcα2-3Galβ1-4GlcNAcβ1-2Manα, which is reported to be the most abundant <i>O</i>-mannosyl glycan on α-DG. Our approach is based on biomimetic stepwise assembly from the reducing end and also gives access to the naturally occurring mono-, di-, and trisaccharide substructures. In addition to the total synthesis, we have developed a “one-pot” enzymatic cascade leading to the rapid synthesis of the target tetrasaccharide. Finally, solid-phase synthesis of the desired glycopeptides directly on a gold microarray platform is described