Chemoenzymatic Synthesis of DSGb5 and Sialylated Globo‐series Glycans

Li, Pei-Jhen; Huang, Szu‐Yu; Chiang, Pei‐Yun; Fan, Chen-Yo; Guo, Li‐Jhen; Wu, Dung-Yeh; Angata, Takashi; Lin, Chun Cheng

doi:10.1002/anie.201903943

Cited by 15 publications

(24 citation statements)

References 32 publications

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“…To gain further insight into the glycan part of Siglec-7 ligands, we sought the sialyltransferase responsible for the biosynthesis of the glycotope recognized by Siglec-7. Siglec-7 preferentially recognizes α2–8–linked oligosialic acids ([Neu5Acα2–8]n; n ≧ 2), disialyl N-acetyllactosamine (Neu5Acα2–3Galβ1–4[Neu5Acα2–6]GlcNAcβ1–), and a terminal tetrasaccharide of α-series gangliosides (Neu5Acα2–3Galβ1–3[Neu5Acα2–6]GalNAcβ1–) ( 46 – 51 ), which are elaborated by ST8Sia and the ST6GalNAc family of sialyltransferases, respectively. Therefore, we analyzed the expression profiles of these sialyltransferases in the JVM-3 cell line; we found that ST8SIA4 and ST6GALNAC4 were highly expressed ( Figure 4A ).…”

Section: Resultsmentioning

confidence: 99%

Molecular Basis and Role of Siglec-7 Ligand Expression on Chronic Lymphocytic Leukemia B Cells

Chang

Liang²,

Lu³

et al. 2022

Front. Immunol.

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Siglec-7 (sialic acid–binding immunoglobulin-like lectin 7) is an immune checkpoint-like glycan recognition protein on natural killer (NK) cells. Cancer cells often upregulate Siglec ligands to subvert immunosurveillance, but the molecular basis of Siglec ligands has been elusive. In this study, we investigated Siglec-7 ligands on chronic lymphocytic leukemia (CLL) B cells. CLL B cells express higher levels of Siglec-7 ligands compared with healthy donor B cells, and enzymatic removal of sialic acids or sialomucins makes them more sensitive to NK cell cytotoxicity. Gene knockout experiments have revealed that the sialyltransferase ST6GalNAc-IV is responsible for the biosynthesis of disialyl-T (Neu5Acα2–3Galβ1–3[Neu5Acα2–6]GalNAcα1–), which is the glycotope recognized by Siglec-7, and that CD162 and CD45 are the major carriers of this glycotope on CLL B cells. Analysis of public transcriptomic datasets indicated that the low expression of GCNT1 (encoding core 2 GlcNAc transferase, an enzyme that competes against ST6GalNAc-IV) and high expression of ST6GALNAC4 (encoding ST6GalNAc-IV) in CLL B cells, together enhancing the expression of the disialyl-T glycotope, are associated with poor patient prognosis. Taken together, our results determined the molecular basis of Siglec-7 ligand overexpression that protects CLL B cells from NK cell cytotoxicity and identified disialyl-T as a potential prognostic marker of CLL.

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Section: Resultsmentioning

confidence: 99%

Molecular Basis and Role of Siglec-7 Ligand Expression on Chronic Lymphocytic Leukemia B Cells

Chang

Liang²,

Lu³

et al. 2022

Front. Immunol.

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“…To facilitate characterization, β-isomer 6 was globally deprotected to give compound 25 . Comparison of the spectroscopic data of the synthetic sample with the 1 H and 13 C NMR spectra reported for authentic chemoenzymatically synthesized Gb5 showed they were identical, establishing the β-anomeric configuration of the penultimate GalNAcβ(1,3)Gal linkage in 6 (Figure S5). The complete deprotection product 22S , obtained from 22 , showed a more downfield-shifted doublet at 5.12 ppm with a J value of 3.6 Hz (Figure S5).…”

Section: Resultsmentioning

confidence: 82%

“…Expanding and/or investigating the functional role of disialyl ganglioside demands the development of a viable approach for accessing well-defined oligosaccharide probe molecules. Recently, modular chemoenzymatic methods for synthesizing DSGb5 glycans, 2 and 3 , containing 6-azidohexyl and 5-aminopentyl aglycones, respectively (Figure ), have been described by us and others. , In both studies, sialyltransferase (ST)-catalyzed enzymatic α(2,3)- and α(2,6)-sialylations at the nonreducing end Gal and inner GalNAc units of a Gb5 pentasaccharide, respectively, were achieved but with low overall enzymatic reaction yields. The intricate oligosaccharide architecture in DSGb5, where two Neu5Acs are bound to the nonreducing end Galβ(1,3)GalNAcβ disaccharide by two different α(2,3)- and α(2,6)-sialyl linkages, further complicates the synthesis of heptasaccharide glycan motif 2 .…”

Section: Introductionmentioning

confidence: 99%

“…In the synthesis of DSGb5, we envisaged that the primary challenge in the developed route would be the stereo- and regioselective α-sialylation to install the α(2,3)- and α(2,6)-sialyl linkages at the nonreducing end Gal and inner GalNAc residues, respectively. Many protected sialic acid donors have beed developed and show excellent stereoselective formation of α-sialyl linkages. − In contrast, chemoenzymatic strategies based on using bacterial STs have greatly facilitated the synthesis of complex sialosides with high regioselectivity and stereoselectivity. , For example, enzymatic α(2,3)-sialylation of Gb5 pentasaccharide effectively generates MSGb5 using cytidine monophosphate-Neu5Ac (CMP-Neu5Ac) and α2,3-ST. , However, few chemical strategies have been developed for the synthesis of Gb5 and its monosialyl congener MSGb5. − Despite continuing advances in glycan synthesis, including some works toward the α(2,6)-sialylation of GalNAc in the Galβ(1,3)GalNAcβ disaccharide unit, the selective α(2,6)-sialylation of the inner GalNAc residue in the Gb5 pentasaccharide template remains unexplored …”

Section: Introductionmentioning

confidence: 99%

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A Modular Chemoenzymatic Synthesis of Disialosyl Globopentaosylceramide (DSGb5Cer) Glycan

Adak

Kuo

et al. 2020

J. Org. Chem.

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The total synthesis of the oligosaccharide moiety of disialosyl globopentaosylceramide (DSGb5 Cer), a dominant ganglioside isolated from malignant renal cell carcinoma tissues, is reported. The synthetic strategy relies on a chemical α(2,6)-sialylation at the internal GalNAc unit of a Gb5 pentasaccharide backbone that furnishes a Neu5Acα(2,6)GalNAc-linked hexasaccharide, suitable for an enzymatic α(2,3)-sialylation of the terminal Gal residue to construct a heptasaccharide glycan. Convergent access to this key α(2,6)-sialylated hexasaccharide was also achieved through a [3+3] glycosylation building upon a Galβ(1,3)[Neu5Acα(2,6)]GalNAc-based trisaccharide donor and a Gb3 acceptor. The synthetic DSGb5 glycan bearing a 6-azidohexyl aglycon at the reducing end could undergo further regioselective functionalization. This approach represents a viable chemoenzymatic method for accessing complex ganglioside glycans and should be useful for the synthesis and biological investigation of DSGb5 derivatives.

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“…Reactions generally proceed under mild, aqueous conditions without the need for toxic and harsh organic reagents. Using bio- and/or chemoenzymatic synthesis tools, several natural and engineered glycan libraries have recently been constructed including asymmetric multi-antennary N -glycans (Wang Z. et al, 2013 ), glycosphingolipid glycans (Yu et al, 2016 ), authentic human type N -glycans (Li L. et al, 2015 ; Hamilton et al, 2017 ), O -mannosyl glycans (Meng et al, 2018 ; Wang S. et al, 2018 ), human milk oligosaccharides (HMOs) (Xiao et al, 2016 ; Prudden et al, 2017 ), and tumor-associated antigens (Li P. J. et al, 2019 ; 'T Hart et al, 2019 ). Similar strategies have been adopted for cell-free enzymatic synthesis of glycolipid libraries including those from bacterial (Glover K. et al, 2005 ), animal (Stubs et al, 2010 ), and human origins (Li S. T. et al, 2019 ).…”

Section: Enzyme-mediated In Vitro Technologies Formentioning

confidence: 99%

Cell-Free Synthetic Glycobiology: Designing and Engineering Glycomolecules Outside of Living Cells

Jaroentomeechai

Taw

et al. 2020

Front. Chem.

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Glycans and glycosylated biomolecules are directly involved in almost every biological process as well as the etiology of most major diseases. Hence, glycoscience knowledge is essential to efforts aimed at addressing fundamental challenges in understanding and improving human health, protecting the environment and enhancing energy security, and developing renewable and sustainable resources that can serve as the source of next-generation materials. While much progress has been made, there remains an urgent need for new tools that can overexpress structurally uniform glycans and glycoconjugates in the quantities needed for characterization and that can be used to mechanistically dissect the enzymatic reactions and multi-enzyme assembly lines that promote their construction. To address this technology gap, cell-free synthetic glycobiology has emerged as a simplified and highly modular framework to investigate, prototype, and engineer pathways for glycan biosynthesis and biomolecule glycosylation outside the confines of living cells. From nucleotide sugars to complex glycoproteins, we summarize here recent efforts that harness the power of cell-free approaches to design, build, test, and utilize glyco-enzyme reaction networks that produce desired glycomolecules in a predictable and controllable manner. We also highlight novel cell-free methods for shedding light on poorly understood aspects of diverse glycosylation processes and engineering these processes toward desired outcomes. Taken together, cell-free synthetic glycobiology represents a promising set of tools and techniques for accelerating basic glycoscience research (e.g., deciphering the “glycan code”) and its application (e.g., biomanufacturing high-value glycomolecules on demand).

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Chemoenzymatic Synthesis of DSGb5 and Sialylated Globo‐series Glycans

Cited by 15 publications

References 32 publications

Molecular Basis and Role of Siglec-7 Ligand Expression on Chronic Lymphocytic Leukemia B Cells

Molecular Basis and Role of Siglec-7 Ligand Expression on Chronic Lymphocytic Leukemia B Cells

A Modular Chemoenzymatic Synthesis of Disialosyl Globopentaosylceramide (DSGb5Cer) Glycan

Cell-Free Synthetic Glycobiology: Designing and Engineering Glycomolecules Outside of Living Cells

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