Syntheses of 6-sulfo sialyl Lewis X glycans corresponding to the L-selectin ligand "sulfoadhesin"

[structure: see text] Divergent syntheses of sulfated sialyl Lewis X oligosaccharides corresponding to the core 1 and core 6 branches of the L-selectin ligand are reported. These synthetic targets incorporate a selectively protected serine residue at the reducing terminus, providing a functional handle for further conjugation.     

A strain-promoted [3 + 2] azide-alkyne cycloaddition for covalent modification of biomolecules in living systems

Selective chemical reactions that are orthogonal to the diverse functionality of biological systems have become important tools in the field of chemical biology. Two notable examples are the Staudinger ligation of azides and phosphines and the Cu(I)-catalyzed [3 + 2] cycloaddition of azides and alkynes ("click chemistry"). The Staudinger ligation has sufficient biocompatibility for performance in living animals but suffers from phosphine oxidation and synthetic challenges. Click chemistry obviates the requirement of phosphines, but the Cu(I) catalyst is toxic to cells, thereby precluding in vivo applications. Here we present a strain-promoted [3 + 2] cycloaddition between cyclooctynes and azides that proceeds under physiological conditions without the need for a catalyst. The utility of the reaction was demonstrated by selective modification of biomolecules in vitro and on living cells, with no apparent toxicity.     

A "traceless" Staudinger ligation for the chemoselective synthesis of amide bonds

[reaction: see text] Here we report a novel modification of our previously reported "Staudinger ligation" that generates an amide bond from an azide and a specifically functionalized phosphine. This method for the selective formation of an amide bond, which does not require the orthogonal protection of distal functional groups, should find general utility in synthetic and biological chemistry.     

Chemical synthesis of lymphotactin: a glycosylated chemokine with a C-terminal mucin-like domain

The synthesis of a 93-residue chemokine, lymphotactin, containing eight sites of O-linked glycosylation, was achieved using the technique of native chemical ligation. A single GalNAc residue was incorporated at each glycosylation site using standard Fmoc-chemistry to achieve the first total synthesis of a mucin-type glycoprotein. Using this approach quantities of homogeneous material were obtained for structural and functional analysis.     

Chemoselective ligation applied to the synthesis of a biantennary N-linked glycoform of CD52

We report here a strategy for the synthesis of N-linked glycopeptide analogues that replace the glycosidic linkages extending from the core pentasaccharide with thioethers amenable to construction by chemoselective ligation. The key building block, a pentasaccharide-Asn analogue containing two thiol residues, was incorporated into CD52 by 9-fluorenylmethoxycarbonyl (Fmoc)-based solid-phase peptide synthesis. An undecasaccharide mimetic was then readily generated by alkylation of this glycopeptide with an N-bromoacetamido trisaccharide. The rapid assembly of a complex type N-linked glycopeptide mimetic was accomplished using this technique.     

An inhibitor of the human UDP-GlcNAc 4-epimerase identified from a uridine-based library: a strategy to inhibit O-linked glycosylation

The biological study of O-linked glycosylation is particularly problematic, as chemical tools to control this modification are lacking. An inhibitor of the UDP-GlcNAc 4-epimerase that synthesizes UDP-GalNAc, the donor initiating O-linked glycosylation, would be a powerful reagent for reversibly inhibiting O-linked glycosylation. We synthesized a 1338 member library of uridine analogs directed to the epimerase by virtue of substrate mimicry. Screening of the library identified an inhibitor with a K(i) value of 11 microM. Tests against related enzymes confirmed the compound's specificity for the UDP-GlcNAc 4-epimerase. Inhibitors of a key step of O-linked glycan biosynthesis can be discovered from a directed library screen. Progeny thereof may be powerful tools for controlling O-linked glycosylation in cells.     

Fusion around the barrier for 7Li+12C

Fusion cross-sections for the ⁷Li + ¹²C reaction have been measured at energies above the Coulomb barrier by the direct detection of evaporation residues. The heavy evaporation residues with energies below 3 MeV could not be separated out from the α-particles in the spectrum and hence their contribution was estimated using statistical model calculations. The present work indicates that suppression of fusion cross-sections due to the breakup of ⁷Li may not be significant for ⁷Li + ¹²C reaction at energies around the barrier.     

Synthetic analogues of glycosylphosphatidylinositol-anchored proteins and their behavior in supported lipid bilayers

Positioned at the C-terminus of many eukaryotic proteins, the glycosylphosphatidylinositol (GPI) anchor is a posttranslational modification that anchors the modified proteins in the outer leaflet of the plasma membrane. GPI-anchored proteins play vital roles in signal transduction, the vertebrate immune response, and the pathobiology of trypanosomal parasites. While many GPI-anchored proteins have been characterized, the biological functions of the GPI anchor have yet to be elucidated at a molecular level. We synthesized a series of GPI-protein analogues bearing modified anchor structures that were designed to dissect the contribution of various glycan components to the GPI-protein's membrane behavior. These anchor analogues were similar in length to native GPI anchors and included mimics of the native structure's three domains. A combination of expressed protein ligation and native chemical ligation was used to attach these analogues to the green fluorescent protein (GFP). These modified GFPs were incorporated in supported lipid bilayers, and their mobilities were analyzed using fluorescence correlation spectroscopy. The data from these experiments suggest that the GPI anchor is more than a simple membrane-anchoring device; it also may prevent transient interactions between the attached protein and the underlying lipid bilayer, thereby permitting rapid diffusion in the bilayer. The ability to generate chemically defined analogues of GPI-anchored proteins is an important step toward elucidating the molecular functions of this interesting post-translational modification.