ArticlePloS one2020
Constructing a human complex type N-linked glycosylation pathway in Kluyveromyces marxianus.
Article in PloS one, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 9 papers.
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Who cites it
9 citing papers in PubMed, 9 citations in OpenAlex.
- Human antibodies as emerging drugs for antimicrobial resistance.Trends in immunology · 2026Review
- Yeast as a Model for Human Disease.International journal of molecular sciences · 2026Review
- Heterologous expressing melittin in a probiotic yeast to evaluate its function for promoting NSC-34 regeneration.Applied microbiology and biotechnology · 2024Article
- Cloning and Optimization of Intracellular Expression of Human Interferon β-1a inAdvanced biomedical research · 2024Article
- RNA polymerase II-driven CRISPR-Cas9 system for efficient non-growth-biased metabolic engineering ofMetabolic engineering communications · 2022Article
- A novel ALG10/TGF-β positive regulatory loop contributes to the stemness of colorectal cancer.Aging · 2022Article
- Strategies to control therapeutic antibody glycosylation during bioprocessing: Synthesis and separation.Biotechnology and bioengineering · 2022Review
- Humanization of Yeasts for Glycan-Type End-Products.Frontiers in microbiology · 2022Review
- The Impact of Glycoengineering on the Endoplasmic Reticulum Quality Control System in Yeasts.Frontiers in molecular biosciences · 2022Review
Corrections and comments
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Authors and funding
7 authors at 4 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Glycosylation can affect various protein properties such as stability, biological activity, and immunogenicity. To produce human therapeutic proteins, a host that can produce glycoproteins with correct glycan structures is required. Microbial expression systems offer economical, rapid and serum-free production and are more amenable to genetic manipulation. In this study, we developed a protocol for CRISPR/Cas9 multiple gene knockouts and knockins in Kluyveromyces marxianus, a probiotic yeast with a rapid growth rate. As hyper-mannosylation is a common problem in yeast, we first knocked out the α-1,3-mannosyltransferase (ALG3) and α-1,6-mannosyltransferase (OCH1) genes to reduce mannosylation. We also knocked out the subunit of the telomeric Ku domain (KU70) to increase the homologous recombination efficiency of K. marxianus. In addition, we knocked in the MdsI (α-1,2-mannosidase) gene to reduce mannosylation and the GnTI (β-1,2-N-acetylglucosaminyltransferase I) and GnTII genes to produce human N-glycan structures. We finally obtained two strains that can produce low amounts of the core N-glycan Man3GlcNAc2 and the human complex N-glycan Man3GlcNAc4, where Man is mannose and GlcNAc is N-acetylglucosamine. This study lays a cornerstone of glycosylation engineering in K. marxianus toward producing human glycoproteins.
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