Engineered N-acetylhexosamine-active enzymes in glycoscience

• Published in: Biochimica et biophysica acta. General subjects Vol. 1861; no. 8; pp. 2070 - 2087
• Main Authors: Slámová, Kristýna, Bojarová, Pavla
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.08.2017
• Subjects: antibodies; beta-N-acetylhexosaminidase; biotechnology; breast milk; Catalysis; catalysts; Chitinase; engineering; genes; glycoproteins; Glycoproteins - biosynthesis; Glycoside Hydrolases - metabolism; Glycosylation; glycosyltransferases; Glycosyltransferases - metabolism; Glycosynthase; Hexosamines - metabolism; low lactose milk; medicine; microorganisms; mutagenesis; mutants; oligosaccharides; Oligosaccharides - biosynthesis; phosphotransferases (kinases); polysaccharides; Protein Engineering; sialic acid; Site-directed mutagenesis; sulfotransferases; Sulfotransferases - metabolism; Transglycosidase; Whole-cell engineering; β-N-acetylhexosaminidase; Chitinase; Glycosynthase; Whole-cell engineering; IgG; UDP-GlcNAc; PAPS; CDP-GlcNAc; Transglycosidase; Fuc; UDP-ManNAc; GlcNAc; SN; UDP-GalNAc; β-N-acetylhexosaminidase; UDP; GH; CAZy; FAD; QM/MM; GT; GalNAc; Gal; Site-directed mutagenesis; O-GlcNAcase; ATP; OGT; PCR
• ISSN: 0304-4165; 1872-8006
• DOI: 10.1016/j.bbagen.2017.03.019

In recent years, enzymes modifying N-acetylhexosamine substrates have emerged in numerous theoretical studies as well as practical applications from biology, biomedicine, and biotechnology. Advanced enzyme engineering techniques converted them into potent synthetic instruments affording a variety of valuable glycosides. This review presents the diversity of engineered enzymes active with N-acetylhexosamine carbohydrates: from popular glycoside hydrolases and glycosyltransferases to less known oxidases, epimerases, kinases, sulfotransferases, and acetylases. Though hydrolases in natura, engineered chitinases, β-N-acetylhexosaminidases, and endo-β-N-acetylglucosaminidases were successfully employed in the synthesis of defined natural and derivatized chitooligomers and in the remodeling of N-glycosylation patterns of therapeutic antibodies. The genes of various N-acetylhexosaminyltransferases were cloned into metabolically engineered microorganisms for producing human milk oligosaccharides, Lewis X structures, and human-like glycoproteins. Moreover, mutant N-acetylhexosamine-active glycosyltransferases were applied, e.g., in the construction of glycomimetics and complex glycostructures, industrial production of low-lactose milk, and metabolic labeling of glycans. In the synthesis of biotechnologically important compounds, several innovative glycoengineered systems are presented for an efficient bioproduction of GlcNAc, UDP-GlcNAc, N-acetylneuraminic acid, and of defined glycosaminoglycans. The above examples demonstrate that engineering of N-acetylhexosamine-active enzymes was able to solve complex issues such as synthesis of tailored human-like glycoproteins or industrial-scale production of desired oligosaccharides. Due to the specific catalytic mechanism, mutagenesis of these catalysts was often realized through rational solutions. Specific N-acetylhexosamine glycosylation is crucial in biological, biomedical and biotechnological applications and a good understanding of its details opens new possibilities in this fast developing area of glycoscience. •Overview of engineered enzymes modifying N-acetylhexosamine glycans is presented.•We show syntheses of humanized glycoproteins, glycomimetics and metabolic labeling.•Engineering of mutant enzymes, recombinant producers and cell factories is shown.•N-acetylhexosamine-active enzymes find use in biotechnology, medicine and research.