Chemoenzymatic Synthesis of a Type 2 Blood Group A Tetrasaccharide and Development of High-throughput Assays Enables a Platform for Screening Blood Group Antigen-cleaving Enzymes

Kwan, David H.; Ernst, Sabrina; Kötzler, Miriam P.; Withers, Stephen G.

doi:10.1093/glycob/cwv031

Cited by 16 publications

(13 citation statements)

References 22 publications

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“…Exchange of the catalytic nucleophilic carboxylate of retaining glycosidases by a small amino acid residue (serine, glycine, or alanine) renders a hydrolytically inactive enzyme that uses fluorinated glycosyl donor substrates for the transfer of the glycosyl moiety to an acceptor molecule (glycosynthase) . The sequential combination of glycosyltransferases and a glycosynthase has recently been demonstrated for the syntheses of a type 2 blood group A tetrasaccharide, glycosphingolipids, and fucosylated xyloglucans . No further examples of the synthesis of complex glycoconjugates by combination of Leloir glycosyltransferases and glycosynthases have been reported so far.…”

Section: Introductionmentioning

confidence: 99%

Combination of Glycosyltransferases and a Glycosynthase in Sequential and One‐Pot Reactions for the Synthesis of Type 1 and Type 2 N‐Acetyllactosamine Oligomers

et al. 2015

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In three words,h ow would you describe your research? Retrosynthetic biocatalysis (of) glycoconjugates. What aspects of this project do you find most exciting? The presented results were only possible by the joint expertise of two research groups and their dedicatedd octoral students within the DFG InternationalR esearch TrainingG roup "SeleCa-Selectivity in Chemo-and Biocatalysis" at RWTH Aachen University and Osaka University.T he Elling group is interested in the synthesis of glycan ligands and glycan inhibitors of galec-tins, al ectin class involved in tumor biology.W ew ere wondering about the possibility to synthesize oligomers with type 1 N-acetyllactosamine (LacNAc) repeats(yellow/blue sugar combination in the cover picture),w hich is known as an unusual glycan structure in somec ancer types. For this goal the expertise of the Pietruszkag roup in the use of glycosynthases was highly valuable. The combination of two glycosyltransferases (blue and red protein structures) and one galactosynthase (yellow protein structure) renders al ibrary of novel poly-LacNAc structures (the yellow/blue and red/blue sugars) in different combinations.T he most exciting aspect is that poly-LacNAc oligomers-not yet found in nature-presenting alternating type 2/type 1L acNAc structures can now be studied for galectin binding and inhibition. What was the inspiration for this cover design? Biocatalysis and especially multi-enzyme systems are av iable alternative fort he synthesis of complex glycan structures. The propera daptation of reaction conditions makes "juggling" with as et of enzymesp ossible. We present novel glycoconju-gates, not yet realized by chemical carbohydrate synthesis. Dedication and Acknowledgements Our paper is dedicated to our colleague Prof. Dr.W olf-Dieter Fessner on the occasion of his 60 th birthday.W ea cknowledge financial supporto fo ur work, as well as the realization of this cover picture, by the DFG within the IRTG 1628 "SeleCa". The front cover artwork for Issue 19/2015 is provided by the groups of Prof. Lothar Elling and Prof. Jçrg Pietruszka (RWTH Aachen University and Heinrich-Heine-Universität Düsseldorf,r espectively). The image shows the combination ("juggling") of two glycosyltransferases(blue and red protein structures) and agalactosynthase (yellow proteinstructure) for the synthesis of ag lycan library. See the Full Paper itself at http://dx.

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Section: Introductionmentioning

confidence: 99%

Combination of Glycosyltransferases and a Glycosynthase in Sequential and One‐Pot Reactions for the Synthesis of Type 1 and Type 2 N‐Acetyllactosamine Oligomers

et al. 2015

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“…Showing the importance of the glycosynthase method in the production of glycoside structures for biological or pharmaceutical research, Kwan et al described the synthesis of the type 2 blood group A oligosaccharide 26 by combining the glycosynthase Abg 2F6 in a reaction sequence with glycosyltranfersases WbgL and BgtA ( Scheme 4 ) [ 33 ]. The enzyme efficiently transferred α- d -galactosyl fluoride ( 2 , αGalF) onto methyl umbelliferone β- N -acetylglucosamine ( 27 , UM-β-GlcNAc) in yields of 84% ( 28 ).…”

Section: Glycoside Syntheses Using Glycosynthase Methodsmentioning

confidence: 99%

Synthesis of Glycosides by Glycosynthases

Hayes

Pietruszka

2017

Molecules

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The many advances in glycoscience have more and more brought to light the crucial role of glycosides and glycoconjugates in biological processes. Their major influence on the functionality and stability of peptides, cell recognition, health and immunity and many other processes throughout biology has increased the demand for simple synthetic methods allowing the defined syntheses of target glycosides. Additional interest in glycoside synthesis has arisen with the prospect of producing sustainable materials from these abundant polymers. Enzymatic synthesis has proven itself to be a promising alternative to the laborious chemical synthesis of glycosides by avoiding the necessity of numerous protecting group strategies. Among the biocatalytic strategies, glycosynthases, genetically engineered glycosidases void of hydrolytic activity, have gained much interest in recent years, enabling not only the selective synthesis of small glycosides and glycoconjugates, but also the production of highly functionalized polysaccharides. This review provides a detailed overview over the glycosylation possibilities of the variety of glycosynthases produced until now, focusing on the transfer of the most common glucosyl-, galactosyl-, xylosyl-, mannosyl-, fucosyl-residues and of whole glycan blocks by the different glycosynthase enzyme variants.

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“…Cleavage of the terminal ␣-GalNAc residue exposed the oligosaccharide to sequential degradation by the ␣-fucosidase, exo-␤-gal, and ␤-hexosaminidase that were included in the assay mixture, resulting in release of the fluorescent methylumbelliferone. Overall, this provided an important demonstration of the potential for improving the activities of these enzymes (53,54). However, the reality that this process would have to be repeated to create good enzymes for Type 3 and 4 linkages and the concern that the presence of a terminal GlcNAc residue could lead to RBC clearance led us to reconsider the wisdom of this specific approach.…”

Section: Figure 3 Overview Of Enzymatic Conversion Of A-and B-antigementioning

confidence: 99%

Toward universal donor blood: Enzymatic conversion of A and B to O type

Rahfeld

Withers

2020

Journal of Biological Chemistry

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Transfusion of blood, or more commonly red blood cells (RBCs), is integral to health care systems worldwide but requires careful matching of blood types to avoid serious adverse consequences. Of the four main blood types, A, B, AB, and O, only O can be given to any patient. This universal donor O-type blood is crucial for emergency situations where time or resources for typing are limited, so it is often in short supply. A and B blood differ from the O type in the presence of an additional sugar antigen (GalNAc and Gal, respectively) on the core H-antigen found on O-type RBCs. Thus, conversion of A, B, and AB RBCs to O-type RBCs should be achievable by removal of that sugar with an appropriate glycosidase. The first demonstration of a B-to-O conversion by Goldstein in 1982 required massive amounts of enzyme but enabled proof-of-principle transfusions without adverse effects in humans. New α-galactosidases and α-N-acetylgalactosaminidases were identified by screening bacterial libraries in 2007, allowing improved conversion of B and the first useful conversions of A-type RBCs, although under constrained conditions. In 2019, screening of a metagenomic library derived from the feces of an AB donor enabled discovery of a significantly more efficient two-enzyme system, involving a GalNAc deacetylase and a galactosaminidase, for A conversion. This promising system works well both in standard conditions and in whole blood. We discuss remaining challenges and opportunities for the use of such enzymes in blood conversion and organ transplantation.

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Chemoenzymatic Synthesis of a Type 2 Blood Group A Tetrasaccharide and Development of High-throughput Assays Enables a Platform for Screening Blood Group Antigen-cleaving Enzymes

Cited by 16 publications

References 22 publications

Combination of Glycosyltransferases and a Glycosynthase in Sequential and One‐Pot Reactions for the Synthesis of Type 1 and Type 2 N‐Acetyllactosamine Oligomers

Combination of Glycosyltransferases and a Glycosynthase in Sequential and One‐Pot Reactions for the Synthesis of Type 1 and Type 2 N‐Acetyllactosamine Oligomers

Synthesis of Glycosides by Glycosynthases

Toward universal donor blood: Enzymatic conversion of A and B to O type

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