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

(2011) TRANSGENIC RESEARCH. 20(5). p.1033-1042
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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 Man(5)GlcNAc(2) 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 Man(8)GlcNAc(2) and Man(7)GlcNAc(2) 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.
Keywords
Recombinant antibody, N-glycosylation, N-ACETYLGLUCOSAMINYLTRANSFERASE-I, Arabidopsis, Molecular farming, HUMAN BETA-1, 4-GALACTOSYLTRANSFERASE, MONOCLONAL-ANTIBODY, MASS-SPECTROMETRY, TRANSGENIC PLANTS, LINKED GLYCANS, GLYCOSYLATION, CELLS, PROTEIN

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Chicago
Henquet, Maurice, Jochem Eigenhuijsen, Thamara Hesselink, Holger Spiegel, Mariëlle Schreuder, Esther van Duijn, Jan Cordewener, Anna Depicker, Alexander van der Krol, and Dirk Bosch. 2011. “Characterization of the Single-chain Fv-Fc Antibody MBP10 Produced in Arabidopsis Alg3 Mutant Seeds.” Transgenic Research 20 (5): 1033–1042.
APA
Henquet, M., Eigenhuijsen, J., Hesselink, T., Spiegel, H., Schreuder, M., van Duijn, E., Cordewener, J., et al. (2011). Characterization of the single-chain Fv-Fc antibody MBP10 produced in Arabidopsis alg3 mutant seeds. TRANSGENIC RESEARCH, 20(5), 1033–1042.
Vancouver
1.
Henquet M, Eigenhuijsen J, Hesselink T, Spiegel H, Schreuder M, van Duijn E, et al. Characterization of the single-chain Fv-Fc antibody MBP10 produced in Arabidopsis alg3 mutant seeds. TRANSGENIC RESEARCH. 2011;20(5):1033–42.
MLA
Henquet, Maurice, Jochem Eigenhuijsen, Thamara Hesselink, et al. “Characterization of the Single-chain Fv-Fc Antibody MBP10 Produced in Arabidopsis Alg3 Mutant Seeds.” TRANSGENIC RESEARCH 20.5 (2011): 1033–1042. Print.
@article{1982170,
  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 Man(5)GlcNAc(2) 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 Man(8)GlcNAc(2) and Man(7)GlcNAc(2) 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.},
  author       = {Henquet, Maurice and Eigenhuijsen, Jochem and Hesselink, Thamara and Spiegel, Holger and Schreuder, Mari{\"e}lle and van Duijn, Esther and Cordewener, Jan and Depicker, Anna and van der Krol, Alexander and Bosch, Dirk},
  issn         = {0962-8819},
  journal      = {TRANSGENIC RESEARCH},
  keyword      = {Recombinant antibody,N-glycosylation,N-ACETYLGLUCOSAMINYLTRANSFERASE-I,Arabidopsis,Molecular farming,HUMAN BETA-1,4-GALACTOSYLTRANSFERASE,MONOCLONAL-ANTIBODY,MASS-SPECTROMETRY,TRANSGENIC PLANTS,LINKED GLYCANS,GLYCOSYLATION,CELLS,PROTEIN},
  language     = {eng},
  number       = {5},
  pages        = {1033--1042},
  title        = {Characterization of the single-chain Fv-Fc antibody MBP10 produced in Arabidopsis alg3 mutant seeds},
  url          = {http://dx.doi.org/10.1007/s11248-010-9475-5},
  volume       = {20},
  year         = {2011},
}

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