Chemoenzymatic Assembly of Mammalian O‐Mannose Glycans

O‐Mannose glycans account up to 30 % of total O‐glycans in the brain. Previous synthesis and functional studies have only focused on the core M3 O‐mannose glycans of α‐dystroglycan, which are a causative factor for various muscular diseases. In this study, a highly efficient chemoenzymatic strategy was developed that enabled the first collective synthesis of 63 core M1 and core M2 O‐mannose glycans. This chemoenzymatic strategy features the gram‐scale chemical synthesis of five judiciously designed core structures, and the diversity‐oriented modification of the core structures with three enzyme modules to provide 58 complex O‐mannose glycans in a linear sequence that does not exceed four steps. The binding profiles of synthetic O‐mannose glycans with a panel of lectins, antibodies, and brain proteins were also explored by using a printed O‐mannose glycan array.     

Divergent Chemoenzymatic Synthesis of Asymmetrical‐Core‐Fucosylated and Core‐Unmodified N‐Glycans

A divergent chemoenzymaytic approach for the preparation of core‐fucosylated and core‐unmodified asymmetrical N‐glycans from a common advances precursor is described. An undecasaccharide was synthesized by sequential chemical glycosylations of an orthogonally protected core fucosylated hexasaccharide that is common to all mammalian core fucosylated N‐glycans. Antennae‐selective enzymatic extension of the undecasaccharide using a panel of glycosyl transferases afforded core fucosylated asymmetrical triantennary N‐glycan isomers, which are potential biomarkers for breast cancer. A unique aspect of our approach is that a fucosidase (FucA1) has been identified that selectively can cleave a core‐fucoside without affecting the fucoside of a sialyl Lewis^X epitope to give easy access to core‐unmodified compounds.     

A Diazido Mannose Analogue as a Chemoenzymatic Synthon for Synthesizing Di‐N‐acetyllegionaminic Acid‐Containing Glycosides

A chemoenzymatic synthon was designed to expand the scope of the chemoenzymatic synthesis of carbohydrates. The synthon was enzymatically converted into carbohydrate analogues, which were readily derivatized chemically to produce the desired targets. The strategy is demonstrated for the synthesis of glycosides containing 7,9‐di‐N‐acetyllegionaminic acid (Leg5,7Ac₂), a bacterial nonulosonic acid (NulO) analogue of sialic acid. A versatile library of α2‐3/6‐linked Leg5,7Ac₂‐glycosides was built by using chemically synthesized 2,4‐diazido‐2,4,6‐trideoxymannose as a chemoenzymatic synthon for highly efficient one‐pot multienzyme (OPME) sialylation followed by downstream chemical conversion of the azido groups into acetamido groups. The syntheses required 10 steps from commercially available d ‐fucose and had an overall yield of 34–52 %, thus representing a significant improvement over previous methods. Free Leg5,7Ac₂ monosaccharide was also synthesized by a sialic acid aldolase‐catalyzed reaction.     

HPLC-Based Automated Synthesis of Glycans in Solution

As the 21^st century unfolds with rapid changes, new challenges in research and development emerge. These new challenges prompted us to repurpose our HPLC−A platform that was previously used in solid phase glycan synthesis to a solution phase batch synthesis described herein. The modular character of HPLC allows for implementing new attachments. To enable sequential synthesis of multiple oligosaccharides with the single press of a button, we supplemented our system with a four-way split valve and an automated fraction collector. This enabled the operator to load all reagents and all reactants in the autosampler, press the button to start the repetitive automation sequence, leave the lab, and upon return find products of multiple reactions ready for purification, analysis, and subsequent application.     

Synthesis of Multiantennary Complex Type N‐Glycans by Use of Modular Building Blocks

A modular set of oligosaccharide building blocks was developed for the synthesis of multiantennary N‐glycans of the complex type, which are commonly found on glycoproteins. The donor building blocks were laid out for the elongation of a core trisaccharide acceptor (β‐mannosyl chitobiose) conveniently protected with a single benzylidene moiety at the β‐mannoside. Through two consecutive regio‐ and stereoselective couplings the donors gave N‐glycans with three to five antennae in high yields. Due to the consistent protection group pattern of the donors the deprotection of the final products can be performed by using a general reaction sequence.     

Stereoselective Synthesis of α‐3‐Deoxy‐D‐manno‐oct‐2‐ulosonic Acid (α‐Kdo) Glycosides Using 5,7‐O‐Di‐tert‐butylsilylene‐Protected Kdo Ethyl Thioglycoside Donors

An efficient methodology for the synthesis of α‐Kdo glycosidic bonds has been developed with 5,7‐O‐di‐tert‐butylsilylene (DTBS) protected Kdo ethyl thioglycosides as glycosyl donors. The approach permits a wide scope of acceptors to be used, thus affording biologically significant Kdo glycosides in good to excellent chemical yields with complete α‐selectivity. The synthetic utility of an orthogonally protected Kdo donor has been demonstrated by concise preparation of two α‐Kdo‐containing oligosaccharides.     

Facile and Versatile Chemoenzymatic Synthesis of Enterobactin Analogues and Applications in Bacterial Detection

Siderophores, such as enterobactin (Ent), are small molecules that can be selectively imported into bacteria along with iron by cognate transporters. Siderophore conjugates are thus a promising strategy for delivering functional reagents into bacteria. In this work, we present an easy‐to‐perform, one‐pot chemoenzymatic synthesis of functionalized monoglucosylated enterobactin (MGE). When functionalized MGE is conjugated to a rhodamine fluorophore, which affords RhB‐Glc‐Ent, it can selectively label Gram‐negative bacteria that utilize Ent, including some E. coli strains and P. aeruginosa. V. cholerae, a bacterium that utilizes linearized Ent, can also be weakly targeted. Moreover, the targeting is effective under iron‐limiting but not iron‐rich conditions. Our results suggest that the RhB‐Glc‐Ent probe is sensitive not only to the bacterial strain but also to the iron condition in the environment.     

A Chemoenzymatic Total Synthesis of the Neurogenic Starfish Ganglioside LLG-3 Using an Engineered and Evolved Synthase

An LLG-3 oligosaccharide-fluoride can be assembled chemoenzymatically and readily coupled with various sphingosines by an engineered endoglycoceramidase glycosynthase. The lyso?ganglioside products are acylated to generate the individual isomers identified in the heterogeneous natural isolates, as well as modified glycosphingolipids.     

化学-酶催化制备天然丹参素

丹参素是中药丹参的水溶性化学成分之一,具有很多潜在的药理活性。 以3,4-二羟基苯甲醛和乙酰甘氨酸为原料,经Erlenmeyer反应并开环得到α-乙酰氨基-β-(3,4-二乙酰氧基苯基)丙烯酸、盐酸水解制得β-(3,4-二羟基苯基)丙酮酸,再经乳酸片球菌1.2696静息细胞生物酶催化还原合成了天然右旋丹参素,总收率69.4%,丹参素对映体过量值97.5%。     

Total Synthesis of the Escherichia coli O111 O‐Specific Polysaccharide Repeating Unit

The first total synthesis of the O‐antigen pentasaccharide repeating unit from Gram‐negative bacteria Escherichia coli O111 was achieved starting from four monosaccharide building blocks. Key to the synthetic approach was a bis‐glycosylation reaction to combine trisaccharide 10 and colitose 5 . The colitose building block ( 5 ) was obtained de novo from non‐carbohydrate precursors. The pentasaccharide was equipped at the reducing end with an amino spacer to provide a handle for subsequent conjugation to a carrier protein in anticipation of immunological studies.     

Machine-Driven Chemoenzymatic Synthesis of Glycopeptide

Historically, researchers have put considerable effort into developing automation systems to prepare natural biopolymers such as peptides and oligonucleotides. The availability of such mature systems has significantly advanced the development of natural science. Over the past twenty years, breakthroughs in automated synthesis of oligosaccharides have also been achieved. A machine-driven platform for glycopeptide synthesis by a reconstructed peptide synthesizer is described. The designed platform is based on the use of an amine-functionalized silica resin to facilitate the chemical synthesis of peptides in organic solvent as well as the enzymatic synthesis of glycan epitopes in the aqueous phase in a single reaction vessel. Both syntheses were performed by a peptide synthesizer in a semiautomated manner.     

Analysis of Density‐Dependent Binding of Glycans by Lectins Using Carbohydrate Microarrays

To investigate the density‐dependent binding of glycans by lectins using carbohydrate microarrays, a number of C‐terminal hydrazide‐conjugated neoglycopeptides with various valences and different spatial arrangements of the sugar ligands were prepared on a solid support. The synthetic strategy includes (1) assembly of alkyne‐linked peptides possessing C‐terminal hydrazide on a solid support, (2) coupling of azide‐linked, unprotected sugars to the alkyne‐linked peptides on the solid support utilizing click chemistry, and (3) release of the neoglycopeptides from the solid support. By using this synthetic methodology, sixty five neoglycopeptides with a valency ranging from 1 to 4 and different spatial arrangements of the carbohydrate ligands were generated. Carbohydrate microarrays were constructed by immobilizing the prepared neoglycopeptides on epoxide‐derivatized glass slides and were used to analyze the density‐dependent binding of glycans by lectins. The results of binding property determinations show that lectin binding is highly dependent on the surface glycan density.     

An Enantio‐ and Diastereoselective Chemoenzymatic Synthesis of α‐Fluoro β‐Hydroxy Carboxylic Esters

The trans‐o‐hydroxybenzylidene pyruvate aldolase‐catalysed reactions between fluoropyruvate and many (hetero)aromatic aldehydes yield aldol adducts without subsequent dehydration. Treatment of the reaction products with hydrogen peroxide yields the corresponding syn‐configured α‐fluoro β‐hydroxy carboxylic acids which have >98 % ee. The overall chemoenzymatic approach, in which fluoropyruvate serves as a fluoroacetate equivalent, may be exploited in the synthesis of polar building blocks and fragments with potential value in drug discovery.     

Current Progress in the Chemoenzymatic Synthesis of Natural Products

Natural products, with their array of structural complexity, diversity, and biological activity, have inspired generations of chemists and driven the advancement of techniques in their total syntheses. The field of natural product synthesis continuously evolves through the development of methodologies to improve stereoselectivity, yield, scalability, substrate scope, late-stage functionalization, and/or enable novel reactions. One of the more interesting and unique techniques to emerge in the last thirty years is the use of chemoenzymatic reactions in the synthesis of natural products. This review highlights some of the recent examples and progress in the chemoenzymatic synthesis of natural products from 2019–2022.     

Modular Chemoenzymatic Synthesis of Terpenes and their Analogues

Non‐natural terpenoids offer potential as pharmaceuticals and agrochemicals. However, their chemical syntheses are often long, complex, and not easily amenable to large‐scale production. Herein, we report a modular chemoenzymatic approach to synthesize terpene analogues from diphosphorylated precursors produced in quantitative yields. Through the addition of prenyl transferases, farnesyl diphosphates, (2E,6E)‐FDP and (2Z,6Z)‐FDP, were isolated in greater than 80 % yields. The synthesis of 14,15‐dimethyl‐FDP, 12‐methyl‐FDP, 12‐hydroxy‐FDP, homo‐FDP, and 15‐methyl‐FDP was also achieved. These modified diphosphates were used with terpene synthases to produce the unnatural sesquiterpenoid semiochemicals (S)‐14,15‐dimethylgermacrene D and (S)‐12‐methylgermacrene D as well as dihydroartemisinic aldehyde. This approach is applicable to the synthesis of many non‐natural terpenoids, offering a scalable route free from repeated chain extensions and capricious chemical phosphorylation reactions.