Combining functional metagenomics and glycoanalytics to identify enzymes that facilitate structural characterization of sulfated N-glycans

Autor:	Mehul B. Ganatra, Erdmann Rapp, Samantha L. Fossa, Udo Reichl, Samanta Cajic, Christopher H. Taron, Léa Chuzel, René Hennig, Madison L. Boisvert
Jazyk:	angličtina
Rok vydání:	2021
Předmět:	Glycan Bioengineering Applied Microbiology and Biotechnology Human microbiome Microbiology Acetylglucosamine Sulfation Capillary electrophoresis Polysaccharides Hexosaminidase chemistry.chemical_classification biology Sulfates Sulfatase Research N-acetylglucosamine-6-sulfate Glycan analysis QR1-502 Enzymes carbohydrates (lipids) Kinetics Enzyme chemistry Biochemistry Metagenomics Pituitary hormones biology.protein Functional metagenomics Glycoanalytics Biotechnology Glycan sulfation
Zdroj:	Microbial Cell Factories, Vol 20, Iss 1, Pp 1-17 (2021) Microbial Cell Factories
ISSN:	1475-2859
Popis:	BackgroundSulfate modification ofN-glycans is important for several biological functions such as clearance of pituitary hormones or immunoregulation. Yet, the prevalence of thisN-glycan modification and its functions remain largely unexplored. Characterization ofN-glycans bearing sulfate modifications is hampered in part by a lack of enzymes that enable site-specific detection ofN-glycan sulfation. In this study, we used functional metagenomic screening to identify enzymes that act upon sulfated N-acetylglucosamine (GlcNAc). Using multiplexed capillary gel electrophoresis with laser-induced fluorescence detection (xCGE-LIF) -based glycoanalysis we proved their ability to act upon GlcNAc-6-SO4onN-glycans.ResultsOur screen identified a sugar-specific sulfatase that specifically removes sulfate from GlcNAc-6-SO4when it is in a terminal position on anN-glycan. Additionally, in the absence of calcium, this sulfatase binds to the sulfated glycan but does not remove the sulfate group, suggesting it could be used for selective isolation of sulfatedN-glycans. Further, we describe isolation of a sulfate-dependent hexosaminidase that removes intact GlcNAc-6-SO4(but not asulfated GlcNAc) from a terminal position onN-glycans. Finally, the use of these enzymes to detect the presence of sulfatedN-glycans by xCGE-LIF is demonstrated.ConclusionThe present study demonstrates the feasibility of using functional metagenomic screening combined with glycoanalytics to discover enzymes that act upon chemical modifications of glycans. The discovered enzymes represent new specificities that can help resolve the presence of GlcNAc-6-SO4inN-glycan structural analyses.
Databáze:	OpenAIRE
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