A critical review of the state-of-the-art evidence in support of the mechanisms of glycosylation reactions is provided. Factors affecting the stability of putative oxocarbenium ions as intermediates at the S1 end of the mechanistic continuum are first surveyed before the evidence, spectroscopic and indirect, for the existence of such species on the time scale of glycosylation reactions is presented. Current models for diastereoselectivity in nucleophilic attack on oxocarbenium ions are then described. Evidence in support of the intermediacy of activated covalent glycosyl donors is reviewed, before the influences of the structure of the nucleophile, of the solvent, of temperature, and of donor-acceptor hydrogen bonding on the mechanism of glycosylation reactions are surveyed. Studies on the kinetics of glycosylation reactions and the use of kinetic isotope effects for the determination of transition-state structure are presented, before computational models are finally surveyed. The review concludes with a critical appraisal of the state of the art.

The synthesis and conformational analysis of a series of phenyl 2,3,6-tri-O-benzyl-β-d-thio galacto- and glucopyranosides and their 6S-deuterio isotopomers, with systematic variation of the protecting group at the 4-position, are described. For the galactopyranosides, replacement of a 4-O-benzyl ether by a 4-O-alkanoyl or aroyl ester results in a small but measurable shift in side chain population away from the trans,gauche conformation and in favor of the gauche,trans conformer. In the glucopyranoside series on the other hand, replacement of a 4-O-benzyl ether by a 4-O-alkanoyl or aroyl ester results in a small but measurable increase in the population of the trans,gauche conformer at the expense of the gauche,gauche conformer. The possible modulating effect of these conformational changes on the well-known changes in the anomeric reactivity of glycosyl donors as a function of protecting group is discussed, raising the possibility that larger changes may be observed at the transition state for glycosylation. A comparable study with a series of ethyl 2,3,4-tri-O-benzyl-β-d-thioglucopyranosides reveals that no significant influence in side chain population is observed on changing the O6 protecting group.

Hexopyranose side chains populate three staggered conformations, whose proportions can be determined from the three sets of ideal limiting J and J coupling constants in combination with the time-averaged experimental coupling constants. Literature values for the limiting coupling constants, obtained by the study of model compounds, the use of the Haasnoot-Altona and related equations, or quantum mechanical computations, can result in computed negative populations of one of the three ideal conformations. Such values arise from errors in the limiting coupling constants and/or from the population of nonideal conformers. We describe the synthesis and analysis of a series of cis- and trans-fused mono-, di-, and trioxabicyclo[4.4.0]octane-like compounds. Correction factors for the application of data from internal models (-CH(OR)-CH(OR)-) to terminal systems (-CH(OR)-CH(OR)) are deduced from comparison of further models, and applied where necessary. Limiting coupling constants so-derived are applied to the side chain conformations of three model hexopyranosides, resulting in calculated conformer populations without negative values. Although, developed primarily for hexopyranose side chains, the limiting coupling constants are suitable, with the correction factors presented, for application to the side chains of higher carbon sugars and to conformation analysis of acyclic diols and their derivatives in a more general sense.

Peptide nucleic acids (PNAs) are promising therapeutic molecules for gene modulation; however, they suffer from poor cell uptake. Delivery of PNAs into cells requires conjugation of the PNA to another large molecule, typically a cell-penetrating peptide or nanoparticle. In this study, we describe a new PNA-based molecule with cyclic tetrahydrofuran (THF) backbone modifications that in some cases considerably improve cell uptake. We refer to these THF-PNA oligomers as thyclotides. With THF groups at every position of the oligomer, the cell uptake of thyclotides targeted to miR-21 is enhanced compared with the corresponding unmodified PNA based on an aminoethylglycine backbone. An optimized thyclotide can efficiently enter cells without the use of cell-penetrating peptides, bind miR-21, its designated microRNA target, decrease expression of miR-21 and increase expression of three downstream targets (PTEN, Cdc25a and KRIT1). Using a plasmid with the PTEN-3′UTR coupled with luciferase, we further confirmed that a miR-21-targeted thyclotide prevents miR-21 from binding to its target RNA. Additionally, the thyclotide shows no cytotoxicity when administered at 200 times its active concentration. We propose that thyclotides be further explored as therapeutic candidates to modulate miRNA levels.