Glycosyl Acceptors Research Articles

Synthesis of 2-Amino-2-deoxy Sugars via Boron-Catalyzed Coupling of Glycosyl Fluorides and Silyl Ether Acceptors.

Although aminosugars are important components in a variety of bioactive molecules, their stereoselective formation is made challenging by the Lewis basic nature of amino substituents. Additionally, the use of N-acyl protecting groups is often problematic due to the competing formation of oxazolines during the glycosylation of 2-aminosugar derivatives. Herein, we report a boron-catalyzed strategy utilizing silyl ether glycosyl acceptors and 2-aminosugar donors that employs the 2,2,2-trichloroethoxycarbonyl (Troc) protecting group for the C2 amino functionality in glycosyl fluorides. This modification allows for operationally simple room-temperature glycosylations and features a rapid reaction profile that addresses some of the limitations in the synthesis of 2-amino-2-deoxy sugar-containing glycosides. Tailoring the order of reactivity of the silyl acceptors enables one-pot iterative glycosylations, thus streamlining the synthesis of complex oligosaccharides while allowing fewer intermediates and purification steps.

Glycoside Phosphorylase Catalyzed Cellulose and β-1,3-Glucan Synthesis Using Chromophoric Glycosyl Acceptors.

Glycoside phosphorylases are enzymes that are frequently used for polysaccharide synthesis. Some of these enzymes have broad substrate specificity, enabling the synthesis of reducing-end-functionalized glucan chains. Here, we explore the potential of glycoside phosphorylases in synthesizing chromophore-conjugated polysaccharides using commercially available chromophoric model compounds as glycosyl acceptors. Specifically, we report cellulose and β-1,3-glucan synthesis using 2-nitrophenyl β-d-glucopyranoside, 4-nitrophenyl β-d-glucopyranoside, and 2-methoxy-4-(2-nitrovinyl)phenyl β-d-glucopyranoside with Clostridium thermocellum cellodextrin phosphorylase and Thermosipho africanus β-1,3-glucan phosphorylase as catalysts. We demonstrate activity for both enzymes with all assayed chromophoric acceptors and report the crystallization-driven precipitation and detailed structural characterization of the synthesized polysaccharides, i.e., their molar mass distributions and various structural parameters, such as morphology, fibril diameter, lamellar thickness, and crystal form. Our results provide insights for the studies of chromophore-conjugated low molecular weight polysaccharides, glycoside phosphorylases, and the hierarchical assembly of crystalline cellulose and β-1,3-glucan.

The impacts of benzoyl and benzyl groups on the non-covalent interactions and electronic properties of glycosyl acceptors

Modulating the reactivity of glycosyl acceptors has become a key in promoting the chemical synthesis of complex glycans. Herein, computational chemistry was employed to explore the impacts of protecting groups on the non-covalent interactions and electronic properties of glycosyl acceptors. Wavefunction analyses showed that substituting benzoyl groups with benzyl groups and introducing a benzyl group to the C2 amine of a D-GlcpNAc residue can eliminate intra-/intermolecular hydrogen bonds, thereby altering the charge distribution significantly. This protecting group-induced charge distribution increases the nucleophilicity of hydroxyl group. This study may contribute to understanding the assistance of computational chemistry in glycan synthesis.

Donor Preactivation-Based Glycan Assembly: from Manual to Automated Synthesis.

Carbohydrates are called the third chain of life. Carbohydrates participate in many important biochemical functions in living species, and the biological information carried by them is several orders of magnitude larger than that of nucleic acids and proteins. However, due to the intrinsic complexity and heterogeneity of carbohydrate structures, furnishing pure and structurally well-defined glycans for functional studies is a formidable task, especially for homogeneous large-size glycans. To address this issue, we have developed a donor preactivation-based one-pot glycosylation strategy enabling multiple sequential glycosylations in a single reaction vessel.The donor preactivation-based one-pot glycosylation refers to the strategy in which the glycosyl donor is activated in the absence of a glycosyl acceptor to generate a reactive intermediate. Subsequently, the glycosyl acceptor with the same anomeric leaving group is added, leading to a glycosyl coupling reaction, which is then iterated to rapidly achieve the desired glycan in the same reactor. The advantages of this strategy include the following: (1) unique chemoselectivity is obtained after preactivation; (2) it is independent of the reactivity of glycosyl donors; (3) multiple-step glycosylations are enabled without the need for intermediate purification; (4) only stoichiometric building blocks are required without complex protecting group manipulations. Using this protocol, a range of glycans including tumor-associated carbohydrate antigens, various glycosaminoglycans, complex N-glycans, and diverse bacterial glycans have been synthesized manually. Gratifyingly, the synthesis of mycobacterial arabinogalactan containing 92 monosaccharide units has been achieved, which created a precedent in the field of polysaccharide synthesis. Recently, the synthesis of a highly branched arabinogalactan from traditional Chinese medicine featuring 140 monosaccharide units has been also accomplished to evaluate its anti-pancreatic-cancer activity. In the spirit of green and sustainable chemistry, this strategy can also be applied to light-driven glycosylation reactions, where either UV or visible light can be used for the activation of glycosyl donors.Automated synthesis is an advanced approach to the construction of complex glycans. Based on the two preactivation modes (general promoter activation mode and light-induced activation mode), a universal and highly efficient automated solution-phase synthesizer was further developed to drive glycan assembly from manual to automated synthesis. Using this synthesizer, a library of oligosaccharides covering various glycoforms and glycosidic linkages was assembled rapidly, either in a general promoter-activation mode or in a light-induced-activation mode. The automated synthesis of a fully protected fondaparinux pentasaccharide was realized on a gram scale. Furthermore, the automated synthesis of large-size polysaccharides was performed, allowing the assembly of arabinans up to an astonishing 1080-mer using the automated multiplicative synthesis strategy, taking glycan synthesis to a new height far beyond the synthesis of nucleic acids (up to 200-mer) and proteins (up to 472-mer).

Harnessing Multistep Chalcogen Bonding Activation in the α-Stereoselective Synthesis of Iminoglycosides.

The use of noncovalent interactions (NCIs) has received significant attention as a pivotal synthetic handle. Recently, the exploitation of unconventional NCIs has gained considerable traction in challenging reaction manifolds such as glycosylation due to their capacity to facilitate entry into difficult-to-access sugars and glycomimetics. While investigations involving oxacyclic pyrano- or furanoside scaffolds are relatively common, methods that allow the selective synthesis of biologically important iminosugars are comparatively rare. Here, we report the capacity of a phosphonochalcogenide (PCH) to catalyze the stereoselective α-iminoglycosylation of iminoglycals with a wide array of glycosyl acceptors with remarkable protecting group tolerance. Mechanistic studies have illuminated the counterintuitive role of the catalyst in serially activating both the glycosyl donor and acceptor in the up/downstream stages of the reaction through chalcogen bonding (ChB). The dynamic interaction of chalcogens with substrates opens up new mechanistic opportunities based on iterative ChB catalyst engagement and disengagement in multiple elementary steps.

GlcNAc-1,6-anhydro-MurNAc Moiety Affords Unusual Glycosyl Acceptor that Terminates Peptidoglycan Elongation.

Peptidoglycan (PG), an essential exoskeletal polymer in bacteria, is a well-known antibiotic target. PG polymerization requires the action of bacterial transglycosylases (TGases), which couple the incoming glycosyl acceptor to the donor. Interfering with the TGase activity can interrupt the PG assembly. Existing TGase inhibitors like moenomycin and Lipid II analogues always occupy the TGase active sites; other strategies to interfere with proper PG elongation have not been widely exploited. Inspired by the natural 1,6-anhydro-MurNAc termini that mark the ends of PG strands in bacteria, we hypothesized that the incorporation of an anhydromuramyl-containing glycosyl acceptor by TGase into the growing PG may effectively inhibit PG elongation. To explore this possibility, we synthesized 4-O-(N-acetyl-β-d-glucosaminyl)-1,6-anhydro-N-acetyl-β-d-muramyl-l-Ala-γ-d-Glu-l-Lys-d-Ala-d-Ala, 1, within 15 steps, and demonstrated that this anhydromuropeptide and its analogue lacking the peptide, 1-deAA, were both utilized by bacterial TGase as noncanonical anhydro glycosyl acceptors in vitro. The incorporation of an anhydromuramyl moiety into PG strands by TGases afforded efficient termination of glycan chain extension. Moreover, the preliminary in vitro studies of 1-deAA against Staphylococcus aureus showed that 1-deAA served as a reasonable antimicrobial adjunct of vancomycin. These insights imply the potential application of such anhydromuropeptides as novel classes of PG-terminating inhibitors, pointing toward novel strategies in antibacterial agent development.

Catalytic Orthogonal Glycosylation Enabled by Enynal‐Derived Copper Carbenes

AbstractHerein, we present an approach for catalytic orthogonal glycosylation utilizing earth‐abundant copper carbenes. This method operates under mild conditions and employs readily accessible starting materials, including benchtop stable enynal‐derived glycosyl donors, synthesized at the gram scale. The reaction accommodates a variety of glycosyl acceptors, including primary, secondary, and tertiary alcohols. The enynal‐derived copper carbenes exhibit remarkable reactivity and selectivity, allowing for the formation of glycosidic linkages with different protecting groups and stereochemical patterns. This approach provides access to both 1,2‐cis‐ and ‐trans‐glycosidic linkages. The product stereoselectivity is independent of the anomeric configuration of the glycosyl donor, which also has orthogonal reactivity to widely used alkynes and thioglycoside donors. An iterative synthesis of a trisaccharide further demonstrates the application of this orthogonal reactivity.

Use of Novel Homochiral Thioureas Camphor Derived as Asymmetric Organocatalysts in the Stereoselective Formation of Glycosidic Bonds.

We synthesized six new camphor-derived homochiral thioureas 1-6, from commercially available (1R)-(-)-camphorquinone. These new compounds 1-6 were evaluated as asymmetric organocatalysts in the stereoselective formation of glycosidic bonds, with 2,3,4,6-tetra-O-benzyl-D-glucopyranosyl and 2,3,4,6-tetra-O-benzyl-D-galactopyranosyl trichloroacetimidates as donors, and several alcohols as glycosyl acceptors, such as methanol, ethanol, 1-propanol, 1-butanol, 1-octanol, iso-propanol, tert-butanol, cyclohexanol, phenol, 1-naphtol, and 2-naphtol. Optimization of the asymmetric glycosylation reaction was achieved by modifying reaction conditions such as solvent, additive, loading of catalyst, temperature, and time of reaction. The best result was obtained with 2,3,4,6-tetra-O-benzyl-D-galactopyranosyl trichloroacetimidates, using 15 mol% of organocatalyst 1, in the presence of 2 equiv of MeOH in solvent-free conditions at room temperature for 1.5 h, affording the glycosidic compound in a 99% yield and 1:73 α:β stereoselectivity; under the same reaction conditions, without using a catalyst, the obtained stereoselectivity was 1:35 α:β. Computational calculations prior to the formation of the products were modeled, using density functional theory, M06-2X/6-31G(d,p) and M06-2X/6-311++G(2d,2p) methods. We observed that the preference for β glycoside formation, through a stereoselective inverted substitution, relies on steric effects and the formation of hydrogen bonds between thiourea 1 and methanol in the complex formed.

Systematic Strategy for the Development of Glycosyltransferase Inhibitors: Diversity-Oriented Synthesis of FUT8 Inhibitors

AbstractGlycans control various biological processes, depending on their structures. Particularly, core fucose, formed by α1,6-fucosyltransferase (FUT8), has a substantial influence on multiple biological processes. In this study, we investigated the development of FUT8 inhibitors with structural elements encompassing both the glycosyl donor (GDP-fucose) and acceptor (N-glycan) of FUT8. To efficiently optimize the structure of FUT8 inhibitors, we employed a strategy involving fragmentation of the target structure, followed by a structure optimization using a diversity-oriented synthesis approach. This study proposes an efficient strategy to accelerate the structural optimization of middle molecules.

Synthesis of Core M3 Matriglycan Constituents via an Additive-Controlled 1,2-cis-Xylopyranosylation with O-Xylosyl Imidates as Donors.

A highly stereoselective strategy for 1,2-cis-xylopyranoside bond formation was established via a preactivation-based, additive-modulated trichloroacetimidate glycosidation strategy. The current protocol is mild, practical, and successful with various xylopyranosyl donors and glycosyl acceptors, including acceptors that are reported to be less reactive due to steric hindrance. The utility of this method was demonstrated with the facile assembly of matriglycan constituent tetra- and hexasaccharides.

Diacetyl strategy for synthesis of NHAc containing glycans: enhancing glycosylation reactivity via diacetyl imide protection.

The presence of NHAc groups in the substrates (both glycosyl donors and acceptors) significantly reduced the reactivity of glycosylation. This decrease was attributed to the NHAc groups forming intermolecular hydrogen bonds by the NHAc groups, thereby reducing molecular mobility. Hence, a diacetyl strategy involving the temporary conversion of NHAc to diacetyl imide (NAc2) was developed for the synthesis of NHAc-containing glycans. This strategy has two significant advantages for oligosaccharide synthesis. The NAc2 protection of NHAc substantially enhances the rate of glycosylation reactions, resulting in improved yields. Moreover, NAc2 can be readily reverted to NHAc by the simple removal of one acetyl group under mild basic conditions, obviating the necessity for treating the polar amino group. We have achieved the efficient synthesis of oligosaccharides containing GlcNHAc and N-glycans containing sialic acid using the diacetyl strategy.

Exploring the Glycosylation Reaction Inside the Resorcin[4]arene Capsule.

In the past decade, there has been an increased interest in applying supramolecular capsule and cage catalysis to the current challenges in synthetic organic chemistry. In this context, we recently reported the resorcin[4]arene capsule-catalyzed conversion of α-glycosyl halides into β-glycosides with high selectivity. Interestingly, this methodology enabled the formation of a wide range of β-pyranosides as well as β-furanosides, although these two donor classes exhibit different reactivities and usually require different reaction conditions and catalysts. Evidence was provided that a proton wire plays a key role in this reaction by enabling dual activation of the glycosyl donor and acceptor. Here, we describe a detailed investigation of several aspects of this reactivity. Besides a mechanistic study, we elucidated the size limitation, the origin of catalytic turnover, and the electrophile scope of nonglycosylic halides. Moreover, a screening of the sensitivity to changes in the reaction conditions provides guidelines to facilitate reproducibility. Furthermore, we demonstrate the compatibility with environmentally benign solvent alternatives, including the renewable solvent limonene.

Flexibility and Function of Distal Substrate-Binding Tryptophans in the Blue Mussel β-Mannanase MeMan5A and Their Role in Hydrolysis and Transglycosylation

β-Mannanases hydrolyze β-mannans, important components of plant and microalgae cell walls. Retaining β-mannanases can also catalyze transglycosylation, forming new β-mannosidic bonds that are applicable for synthesis. This study focused on the blue mussel (Mytilus edulis) GH5_10 β-mannanase MeMan5A, which contains two semi-conserved tryptophans (W240 and W281) in the distal subsite +2 of its active site cleft. Variants of MeMan5A were generated by replacing one or both tryptophans with alanines. The substitutions reduced the enzyme’s catalytic efficiency (kcat/Km using galactomannan) by three-fold (W281A), five-fold (W240A), or 20-fold (W240A/W281A). Productive binding modes were analyzed by 18O labeling of hydrolysis products and mass spectrometry. Results show that the substitution of both tryptophans was required to shift away from the dominant binding mode of mannopentaose (spanning subsites −3 to +2), suggesting that both tryptophans contribute to glycan binding. NMR spectroscopy and molecular dynamics simulations were conducted to analyze protein flexibility and glycan binding. We suggest that W240 is rigid and contributes to +2 subsite mannosyl specificity, while W281 is flexible, which enables stacking interactions in the +2 subsite by loop movement to facilitate binding. The substitutions significantly reduced or eliminated transglycosylation with saccharides as glycosyl acceptors but had no significant effect on reactions with alcohols.

Structure, catalysis, chitin transport, and selective inhibition of chitin synthase

Chitin is one of the most abundant natural biopolymers and serves as a critical structural component of extracellular matrices, including fungal cell walls and insect exoskeletons. As a linear polymer of β-(1,4)-linked N-acetylglucosamine, chitin is synthesized by chitin synthases, which are recognized as targets for antifungal and anti-insect drugs. In this study, we determine seven different cryo-electron microscopy structures of a Saccharomyces cerevisiae chitin synthase in the absence and presence of glycosyl donor, acceptor, product, or peptidyl nucleoside inhibitors. Combined with functional analyses, these structures show how the donor and acceptor substrates bind in the active site, how substrate hydrolysis drives self-priming, how a chitin-conducting transmembrane channel opens, and how peptidyl nucleoside inhibitors inhibit chitin synthase. Our work provides a structural basis for understanding the function and inhibition of chitin synthase.

L‐Proline Derived Thioamide Small Organic Molecule for the α‐Stereoselective Synthesis of 2‐Deoxyglycosides

AbstractL‐Prolinethioamide catalyzes direct stereoselective glycosylation of glycal donors to access 2‐deoxyglycosides under moderate reaction conditions. This proposed glycosylation protocol produced the desired products in up to 88% yield and shows greater tolerance of glycosyl acceptors and a broad substrate scope with better stereoselectivity. This method is also applicable for less nucleophilic acceptors such as phenol. In addition, 1,1′‐linked trehalose‐type analogues can also be accessed through this methodology. Further, mechanistic studies suggest that, in this glycosylation, L‐prolinethioamide operates via Brønsted acid/base catalysis.

Activation of thioglycosides under mild alkylation conditions

Reported herein is the development of a novel method for the activation of thioglycosides and thioimidates using benzyl trichloroacetimidate in the presence of catalytic triflic acid. Excellent yields have been achieved with reactive substrates, whereas efficiency of reactions with unreactive glycosyl donors and/or acceptors was modest.

Computational study of chemical phenol glycosylation mechanism in the gas phase for modeling direct glycoconjugate formation in raw plant material

Glycosylation in a broad sense is commonly considered a chemical reaction between a carbohydrate and another molecule with any functional group which can act as a glycosyl acceptor forming covalent bond. Such modification of small natural biologically active molecules (e.g., flavonols) is of significant importance for drug delivery and dietary supplements due to the increase in their bioavailability. Nevertheless, classical chemical synthesis is complicated by the oxidation of flavonols and requires multiple stages to prevent its degradation. Thus, the idea of direct solid-state chemical synthesis in raw plant material arises. To evaluate such a possibility, we provide a computational gas phase study of this process based on the model system of reaction between glucose and phenol. This investigation is based on the assumption that water and flavonol (here phenol as a model) molecules are deficient in raw plant material and do not form liquid phase, whereas dielectric constant (Ɛ = 2–5) is close to 1 in vacuo, and absence of regular-crystals structure. The reaction mechanism and particular path of phenol direct chemical glycosylation are calculated as well as all intermediate products, transition states, and energy barriers tacking into account molecular symmetry. This work proves a concept of the possibility of such reaction in the raw plant material, including flavonol direct glycosylation using mechanochemistry.

Directed SN2 Glycosylation Employing an Amide-Functionalized 1-Naphthoate Platform Featuring a Selectivity-Safeguarding Mechanism.

This work implements a catalytic SN2 glycosylation by employing an amide-functionalized 1-naphthoate platform as a latent glycosyl leaving group. Upon gold-catalyzed activation, the amide group enables the SN2 process by directing the attack of the glycosyl acceptor via H-bonding interaction, which results in stereoinversion at the anomeric center. Unique in this approach is that the amide group also enables a novel safeguarding mechanism by trapping oxocarbenium intermediates and, hence, minimizing stereorandom SN1 processes. The strategy is applicable to the synthesis of a broad range of glycosides with high to excellent levels of stereoinversion from anomerically pure/enriched glycosyl donors. These reactions are generally high-yielding, and their applications in the synthesis of challenging 1,2-cis-linkage-rich oligosaccharides are demonstrated.

BF3 ⋅ CH3CN Promotes a Formation of Aryl α‐O‐Glycosides from Pyranoses: A Facile Approach to Oligosaccharide EPS‐G7

AbstractA BF3 ⋅ CH3CN‐promoted synthesis of aryl α‐O‐glycosides from readily available glycosyl donors was developed. This reaction could proceed well on a gram‐scale under mild condition with α/β ratio up to 95 : 5. Moreover, a variety of the glycosyl donors including acetylated/benzoylated pyranoses, di‐ and trisaccharide derivatives as well as glycosyl acceptors from substituted phenols, sterically hindered aliphatic alcohols and carboxylic acid, were well tolerant. Importantly, the potential of this strategy was highlighted with a successful gram‐scale synthesis of oligosaccharide EPS‐G7, which is a commercial detection reagent for quantitative determination of α‐amylase.

Synthesis of Unprecedented α/β-Alternate (1→4)-Glucans via Stereoselective Iterative Glycosylation.

Glucans are major biomaterials on the earth, with α-(1→4)-glucans (i. e., amylose) and β-(1→4)-glucans (i. e., cellulose) being the most abundant ones, which are relevant to energy storage and structural function, respectively. Interestingly, (1→4)-glucans with alternate α/β-linkages, namely herewith amycellulose, have never been disclosed in nature. Here we report a robust glycosylation protocol for the stereoselective construction of the 1,2-cis-α- and 1,2-trans-β-glucosidic linkages, which employs an optimal combination of glycosyl N-phenyltrifluoroacetimidates as donors, TMSNTf2 as promoter, CH2 Cl2 /nitrile or CH2 Cl2 /THF as solvents. A broad substrate scope has been demonstrated by coupling five imidate donors with eight glycosyl acceptors, in which most of the glycosylations lead to high yield and exclusively 1,2-cis-α- or 1,2-trans-β-selectivity. Applying this glycosylation protocol and with an iterative manner, the unprecedented α/β-alternate (1→4)-glucans up to a 16-mer have been synthesized. Differently from amylose, that adopts a compact helicoidal arrangement, the synthetic amycellulose features an extended ribbon-like conformation, comparable to the extended shape of cellulose.