Production of galactosylated complex-type N-glycans in glycoengineered Saccharomyces cerevisiae
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A1 Alkuperäisartikkeli tieteellisessä aikakauslehdessä
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15
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Applied Microbiology & Biotechnology, Volume 106, issue 1, pp. 301-315
Abstract
N-glycosylation is an important posttranslational modification affecting the properties and quality of therapeutic proteins. Glycoengineering in yeast aims to produce proteins carrying human-compatible glycosylation, enabling the production of therapeutic proteins in yeasts. In this work, we demonstrate further development and characterization of a glycoengineering strategy in a Saccharomyces cerevisiae Δalg3 Δalg11 strain where a truncated Man3GlcNAc2 glycan precursor is formed due to a disrupted lipid-linked oligosaccharide synthesis pathway. We produced galactosylated complex-type and hybrid-like N-glycans by expressing a human galactosyltransferase fusion protein both with and without a UDP-glucose 4-epimerase domain from Schizosaccharomyces pombe. Our results showed that the presence of the UDP-glucose 4-epimerase domain was beneficial for the production of digalactosylated complex-type glycans also when extracellular galactose was supplied, suggesting that the positive impact of the UDP-glucose 4-epimerase domain on the galactosylation process can be linked to other processes than its catalytic activity. Moreover, optimization of the expression of human GlcNAc transferases I and II and supplementation of glucosamine in the growth medium increased the formation of galactosylated complex-type glycans. Additionally, we provide further characterization of the interfering mannosylation taking place in the glycoengineered yeast strain.Description
Lisään tiedoston kun VoR julkaistaan. Funding Information: Open Access funding provided by Aalto University. This work was supported by Aalto University School of Chemical Engineering and the Finnish Foundation for Technology Promotion to MP and by Academy of Finland (grant number 298476) to AF. Publisher Copyright: © 2021, The Author(s).
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Piirainen, M A, Salminen, H & Frey, A D 2022, 'Production of galactosylated complex-type N-glycans in glycoengineered Saccharomyces cerevisiae', Applied Microbiology & Biotechnology, vol. 106, no. 1, pp. 301-315. https://doi.org/10.1007/s00253-021-11727-8