Engineering of CHO Cells for the Production of Recombinant Glycoprotein Vaccines with Xylosylated N-glycans.

Grit Sandig,Hans Von Horsten,Volker Sandig,Stephan Hinderlich,Marcus Frohme,Véronique Blanchard,Lars Radke,Christoph Giese

doi:10.3390/bioengineering4020038

Abstract

Xylose is a general component of O-glycans in mammals. Core-xylosylation of N-glycans is only found in plants and helminth. Consequently, xylosylated N-glycans cause immunological response in humans. We have used the F-protein of the human respiratory syncytial virus (RSV), one of the main causes of respiratory tract infection in infants and elderly, as a model protein for vaccination. The RSV-F protein was expressed in CHO-DG44 cells, which were further modified by co-expression of β1,2-xylosyltransferase from Nicotiana tabacum. Xylosylation of RSV-F N-glycans was shown by monosaccharide analysis and MALDI-TOF mass spectrometry. In immunogenic studies with a human artificial lymph node model, the engineered RSV-F protein revealed improved vaccination efficacy.

Highlights

Glycosylation is one of the most important post-translational modifications
The respiratory syncytial virus (RSV)-F protein was expressed in CHO-DG44 cells, which were further modified by co-expression of β1,2-xylosyltransferase from Nicotiana tabacum
RreSsVs-iFonp,rointedinicdaetirnivgedsufrcocmessful co-expthreessesicolnonoefs bwoatshlapbreolteedin“sR.SRVS-FVX-Fylp+r”o. tein derived from these clones was labeled “RSV-F Xyl+”

Summary

Introduction

Glycosylation is one of the most important post-translational modifications. Proteins can be decorated with glycans, which are linked either with asparagine residues via an N-glycosidic linkage (N-glycans) or with serine or threonine residues via an O-glycosidic linkage (O-glycans) [1]. The majority of N- and O-glycans consists of six different monosaccharides, namely mannose, galactose, N-acetylglucosamine (GlcNAc), N-acetylgalactosamine (GalNAc), fucose and sialic acids. Xylose is a rare monosaccharide component of O-glycans [2], and it is part of the linker between glycosaminoglycans and proteins [3]. Pentose, together with glucuronic acid, is one of the two monosaccharide components of a heteropolymer linked to α-dystroglycan in muscle cells [4], where it mediates its interaction with the extracellular matrix component laminin. In N-glycans xylose is only found in plants and helminth [5,6]. There it is usually β1,2-linked to the central mannose residue of the N-glycan core

Methods

Results

Conclusion