Enzymatic Cascades for Tailored 13C6 and 15N Enriched Human Milk Oligosaccharides

Fischöder, Thomas; Cajic, Samanta; Grote, Valerian; Heinzler, Raphael; Reichl, Udo; Franzreb, Matthias; Rapp, Erdmann; Elling, Lothar

doi:10.3390/molecules24193482

Cited by 9 publications

(5 citation statements)

References 97 publications

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“…MALDI-TOF-MS analysis of released N-glycans was performed as described previously (Selman et al, 2011;Fischöder et al, 2019). Briefly, 0.9 cm cotton rope was used for Cotton HILIC SPE.…”

Section: Matrix-assisted Laser Desorption/ionization Time-of-flight Mass Spectrometry Based N-glycan Analysismentioning

confidence: 99%

Cell-Free Glycoengineering of the Recombinant SARS-CoV-2 Spike Glycoprotein

Ruhnau

Grote

Juarez-Osorio

et al. 2021

Front. Bioeng. Biotechnol.

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The baculovirus-insect cell expression system is readily utilized to produce viral glycoproteins for research as well as for subunit vaccines and vaccine candidates, for instance against SARS-CoV-2 infections. However, the glycoforms of recombinant proteins derived from this expression system are inherently different from mammalian cell-derived glycoforms with mainly complex-type N-glycans attached, and the impact of these differences in protein glycosylation on the immunogenicity is severely under investigated. This applies also to the SARS-CoV-2 spike glycoprotein, which is the antigen target of all licensed vaccines and vaccine candidates including virus like particles and subunit vaccines that are variants of the spike protein. Here, we expressed the transmembrane-deleted human β-1,2 N-acetlyglucosamintransferases I and II (MGAT1ΔTM and MGAT2ΔTM) and the β-1,4-galactosyltransferase (GalTΔTM) in E. coli to in-vitro remodel the N-glycans of a recombinant SARS-CoV-2 spike glycoprotein derived from insect cells. In a cell-free sequential one-pot reaction, fucosylated and afucosylated paucimannose-type N-glycans were converted to complex-type galactosylated N-glycans. In the future, this in-vitro glycoengineering approach can be used to efficiently generate a wide range of N-glycans on antigens considered as vaccine candidates for animal trials and preclinical testing to better characterize the impact of N-glycosylation on immunity and to improve the efficacy of protein subunit vaccines.

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“…MALDI-TOF-MS analysis of released N-glycans was performed as described previously (Selman et al, 2011;Fischöder et al, 2019). Briefly, 0.9 cm cotton rope was used for Cotton HILIC SPE.…”

Section: Matrix-assisted Laser Desorption/ionization Time-of-flight Mass Spectrometry Based N-glycan Analysismentioning

confidence: 99%

Cell-Free Glycoengineering of the Recombinant SARS-CoV-2 Spike Glycoprotein

Ruhnau

Grote

Juarez-Osorio

et al. 2021

Front. Bioeng. Biotechnol.

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“…Kinase-dependent pathways have been reported for the synthesis of various nucleotide sugars including UDP-Gal and UDP-GalNAc [177], GDP-mannose [180], GDP-fucose [181], as well as nucleotide sugars with unnatural sugar moieties [167,[182][183][184]. GDP-mannose has been synthesized also via a kinase-independent route from mannose 1-phosphate produced in two steps from sucrose [185].…”

Section: Enzymatic Synthesis Of Nucleotide Sugarsmentioning

confidence: 99%

Nucleotide Sugars in Chemistry and Biology

Mikkola

2020

Molecules

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Nucleotide sugars have essential roles in every living creature. They are the building blocks of the biosynthesis of carbohydrates and their conjugates. They are involved in processes that are targets for drug development, and their analogs are potential inhibitors of these processes. Drug development requires efficient methods for the synthesis of oligosaccharides and nucleotide sugar building blocks as well as of modified structures as potential inhibitors. It requires also understanding the details of biological and chemical processes as well as the reactivity and reactions under different conditions. This article addresses all these issues by giving a broad overview on nucleotide sugars in biological and chemical reactions. As the background for the topic, glycosylation reactions in mammalian and bacterial cells are briefly discussed. In the following sections, structures and biosynthetic routes for nucleotide sugars, as well as the mechanisms of action of nucleotide sugar-utilizing enzymes, are discussed. Chemical topics include the reactivity and chemical synthesis methods. Finally, the enzymatic in vitro synthesis of nucleotide sugars and the utilization of enzyme cascades in the synthesis of nucleotide sugars and oligosaccharides are briefly discussed.

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“…However, diluted CO 2 streams are no promising entry point for a closed-loop value chain. Therefore, a direct chemical recycling should be considered [1,2]. In this work, the chemical recycling of PHB and PLA over supported catalysts, such as Ru/CeO 2 , under hydrogen atmosphere towards the respective monomers is demonstrated with high yields of 79 % and 94 %.…”

Section: P402mentioning

confidence: 96%

“…Different structures are built stepwise by reactions catalyzed by different glycosyltransferases. The combination of multiple enzymes and sugar acceptor/donors leads to the production of different glycan structures in cascades [1]. Compartmented reactors involving immo-bilized enzymes allow sequential glycosylation and product recovery [2].…”

Section: P403mentioning

confidence: 99%