Glycoengineering of the methylotrophic yeast Hansenula polymorpha for the production of glycoproteins with trimannosyl core N-glycan by blocking core oligosaccharide assembly

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Title
Glycoengineering of the methylotrophic yeast Hansenula polymorpha for the production of glycoproteins with trimannosyl core N-glycan by blocking core oligosaccharide assembly
Author(s)
Doo-Byoung Oh; J S Park; Moo Woong Kim; Seon Ah Cheon; Eun Jung Kim; Hye Yun Moon; Oh Suk Kwon; Sang Ki Rhee; Hyun Ah Kang
Bibliographic Citation
Biotechnology Journal, vol. 3, no. 5, pp. 659-668
Publication Year
2008
Abstract
The initial lipid-linked oligosaccharide Glc3Man9GlcNAc2-dolichyl pyrophosphate (Dol-PP) for N-glycan is synthesized and assembled at the membrane of the endoplasmic reticulum (ER) and subsequently transferred to a nascent polypeptide by the oligosaccharide transferase complex. We have identified an ALG3 homolog (HpALG3) coding for a dolichyl-phosphate-mannose dependent α-1,3-mannosyltransferase in the methylotrophic yeast Hansenula polymorpha. The detailed analysis of glycan structure by linkage-specific mannosidase digestion showed that HpALG3 is responsible for the conversion of MAn5GlcNAc2-Dol-PP to Man6GlcNAc2-Dol-PP, the first step to attach a mannose to the lipid-linked oligosaccharide in the ER. The N-glycosylation pathway of H. polymorpha has been remodeled by deleting the HpALG3 gene in the Hpoch1 null mutant strain blocked in the yeast-specific outer mannose chain synthesis and by introducing an ER-targeted Aspergillus saitoi α-1,2-mannosidase gene. This glycoengineered H. polymorpha strain produced glycoproteins mainly containing trimannosyl core N-glycan (Man3GlcNAc2), which is the common core backbone of various human-type N-glycans. The results demonstrate the high potential of H. polymorpha to be developed as an efficient expression system for the production of glycoproteins with humanized glycans.
Keyword
ALG3GlycoengineeringHansenula polymorphaTrimannosyl core N-glycan
ISSN
1860-6768
Publisher
Wiley
DOI
http://dx.doi.org/10.1002/biot.200700252
Type
Article
Appears in Collections:
Division of Bio Technology Innovation > SME Support Center > 1. Journal Articles
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