Characterization of the single-chain Fv-Fc antibody MBP10 produced in Arabidopsis alg3 mutant seeds

M.G.L. Henquet, J. Eigenhuijsen, T. Hesselink, H. Spiegel, M.E.L. Schreuder, E. van Duijn, J.H.G. Cordewener, A. Depicker, A.R. van der Krol, H.J. Bosch

Research output: Contribution to journalArticleAcademicpeer-review

6 Citations (Scopus)

Abstract

ER resident glycoproteins, including ectopically expressed recombinant glycoproteins, carry so-called high-mannose type N-glycans, which can be at different stages of processing. The presence of heterogeneous high-mannose type glycans on ER-retained therapeutic proteins is undesirable for specific therapeutic applications. Previously, we described an Arabidopsis alg3-2 glycosylation mutant in which aberrant Man5GlcNAc2 mannose type N-glycans are transferred to proteins. Here we show that the alg3-2 mutation reduces the N-glycan heterogeneity on ER resident glycoproteins in seeds. We compared the properties of a scFv-Fc, with a KDEL ER retention tag (MBP10) that was expressed in seeds of wild type and alg3-2 plants. N-glycans on these antibodies from mutant seeds were predominantly of the intermediate Man5GlcNAc2 compared to Man8GlcNAc2 and Man7GlcNAc2 isoforms on MBP10 from wild-type seeds. The presence of aberrant N-glycans on MBP10 did not seem to affect MBP10 dimerisation nor binding of MBP10 to its antigen. In alg3-2 the fraction of underglycosylated MBP10 protein forms was higher than in wild type. Interestingly, the expression of MBP10 resulted also in underglycosylation of other, endogenous glycoproteins
Original languageEnglish
Pages (from-to)1033-1042
JournalTransgenic Research
Volume20
Issue number5
DOIs
Publication statusPublished - 2011

Keywords

  • n-acetylglucosaminyltransferase-i
  • human beta-1,4-galactosyltransferase
  • monoclonal-antibody
  • mass-spectrometry
  • transgenic plants
  • linked glycans
  • glycosylation
  • cells
  • protein

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