Off‐target glycans encountered along the ...
Type de document :
Article dans une revue scientifique
DOI :
URL permanente :
Titre :
Off‐target glycans encountered along the synthetic biology route toward humanized N‐glycans in Pichia pastoris
Auteur(s) :
Laukens, Bram [Auteur]
Universiteit Gent = Ghent University [UGENT]
Jacobs, Pieter P. [Auteur]
Universiteit Gent = Ghent University [UGENT]
Geysens, Katelijne [Auteur]
Universiteit Gent = Ghent University [UGENT]
Martins, Jose [Auteur]
Universiteit Gent = Ghent University [UGENT]
De Wachter, Charlot [Auteur]
Universiteit Gent = Ghent University [UGENT]
Ameloot, Paul [Auteur]
Universiteit Gent = Ghent University [UGENT]
Morelle, Willy [Auteur]
Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF]
Haustraete, Jurgen [Auteur]
Vlaams Instituut voor Biotechnologie [Ghent, Belgique] [VIB]
Renauld, Jean‐Christophe [Auteur]
Université Catholique de Louvain = Catholic University of Louvain [UCL]
Samyn, Bart [Auteur]
Universiteit Gent = Ghent University [UGENT]
Contreras, Roland [Auteur]
Universiteit Gent = Ghent University [UGENT]
Devos, Simon [Auteur]
Vlaams Instituut voor Biotechnologie [Ghent, Belgique] [VIB]
Universiteit Gent = Ghent University [UGENT]
Callewaert, Nico [Auteur]
Vlaams Instituut voor Biotechnologie [Ghent, Belgique] [VIB]
Universiteit Gent = Ghent University [UGENT]
Universiteit Gent = Ghent University [UGENT]
Jacobs, Pieter P. [Auteur]
Universiteit Gent = Ghent University [UGENT]
Geysens, Katelijne [Auteur]
Universiteit Gent = Ghent University [UGENT]
Martins, Jose [Auteur]
Universiteit Gent = Ghent University [UGENT]
De Wachter, Charlot [Auteur]
Universiteit Gent = Ghent University [UGENT]
Ameloot, Paul [Auteur]
Universiteit Gent = Ghent University [UGENT]
Morelle, Willy [Auteur]
Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF]
Haustraete, Jurgen [Auteur]
Vlaams Instituut voor Biotechnologie [Ghent, Belgique] [VIB]
Renauld, Jean‐Christophe [Auteur]
Université Catholique de Louvain = Catholic University of Louvain [UCL]
Samyn, Bart [Auteur]
Universiteit Gent = Ghent University [UGENT]
Contreras, Roland [Auteur]
Universiteit Gent = Ghent University [UGENT]
Devos, Simon [Auteur]
Vlaams Instituut voor Biotechnologie [Ghent, Belgique] [VIB]
Universiteit Gent = Ghent University [UGENT]
Callewaert, Nico [Auteur]
Vlaams Instituut voor Biotechnologie [Ghent, Belgique] [VIB]
Universiteit Gent = Ghent University [UGENT]
Titre de la revue :
Biotechnology and bioengineering
Nom court de la revue :
Biotechnology and Bioengineering
Numéro :
117
Pagination :
2479-2488
Éditeur :
Wiley
Date de publication :
2020-05-26
Mot(s)-clé(s) en anglais :
GlycoSwitch
N‐glycan engineering
Pichia pastoris
synthetic biology
N‐glycan engineering
Pichia pastoris
synthetic biology
Discipline(s) HAL :
Sciences du Vivant [q-bio]
Résumé en anglais : [en]
The glycosylation pathways of several eukaryotic protein expression hosts are being engineered to enable the production of therapeutic glycoproteins with humanized application‐customized glycan structures. In several ...
Lire la suite >The glycosylation pathways of several eukaryotic protein expression hosts are being engineered to enable the production of therapeutic glycoproteins with humanized application‐customized glycan structures. In several expression hosts, this has been quite successful, but one caveat is that the new N‐glycan structures inadvertently might be substrates for one or more of the multitude of endogenous glycosyltransferases in such heterologous background. This then results in the formation of novel, undesired glycan structures, which often remain insufficiently characterized. When expressing mouse interleukin‐22 in a Pichia pastoris (syn. Komagataella phaffii) GlycoSwitchM5 strain, which had been optimized to produce Man5GlcNAc2N‐glycans, glycan profiling revealed two major species: Man5GlcNAc2 and an unexpected, partially α‐mannosidase‐resistant structure. A detailed structural analysis using exoglycosidase sequencing, mass spectrometry, linkage analysis, and nuclear magnetic resonance revealed that this novel glycan was Man5GlcNAc2 modified with a Glcα‐1,2‐Manβ‐1,2‐Manβ‐1,3‐Glcα‐1,3‐R tetrasaccharide. Expression of a Golgi‐targeted GlcNAc transferase‐I strongly inhibited the formation of this novel modification, resulting in more homogeneous modification with the targeted GlcNAcMan5GlcNAc2 structure. Our findings reinforce accumulating evidence that robustly customizing the N‐glycosylation pathway in P. pastoris to produce particular human‐type structures is still an incompletely solved synthetic biology challenge, which will require further innovation to enable safe glycoprotein pharmaceutical production.Lire moins >
Lire la suite >The glycosylation pathways of several eukaryotic protein expression hosts are being engineered to enable the production of therapeutic glycoproteins with humanized application‐customized glycan structures. In several expression hosts, this has been quite successful, but one caveat is that the new N‐glycan structures inadvertently might be substrates for one or more of the multitude of endogenous glycosyltransferases in such heterologous background. This then results in the formation of novel, undesired glycan structures, which often remain insufficiently characterized. When expressing mouse interleukin‐22 in a Pichia pastoris (syn. Komagataella phaffii) GlycoSwitchM5 strain, which had been optimized to produce Man5GlcNAc2N‐glycans, glycan profiling revealed two major species: Man5GlcNAc2 and an unexpected, partially α‐mannosidase‐resistant structure. A detailed structural analysis using exoglycosidase sequencing, mass spectrometry, linkage analysis, and nuclear magnetic resonance revealed that this novel glycan was Man5GlcNAc2 modified with a Glcα‐1,2‐Manβ‐1,2‐Manβ‐1,3‐Glcα‐1,3‐R tetrasaccharide. Expression of a Golgi‐targeted GlcNAc transferase‐I strongly inhibited the formation of this novel modification, resulting in more homogeneous modification with the targeted GlcNAcMan5GlcNAc2 structure. Our findings reinforce accumulating evidence that robustly customizing the N‐glycosylation pathway in P. pastoris to produce particular human‐type structures is still an incompletely solved synthetic biology challenge, which will require further innovation to enable safe glycoprotein pharmaceutical production.Lire moins >
Langue :
Anglais
Audience :
Non spécifiée
Projet Européen :
Établissement(s) :
Université de Lille
CNRS
CNRS
Équipe(s) de recherche :
Mécanismes moléculaires de la N-glycosylation et pathologies associées
Date de dépôt :
2021-03-23T15:10:52Z
2021-03-24T09:28:04Z
2021-03-24T09:28:04Z