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Establishment of a tobacco BY2 cell line devoid of plant-specific xylose and fucose as a platform for the production of biotherapeutic proteins.
Hanania, Uri; Ariel, Tami; Tekoah, Yoram; Fux, Liat; Sheva, Maor; Gubbay, Yehuda; Weiss, Mara; Oz, Dina; Azulay, Yaniv; Turbovski, Albina; Forster, Yehava; Shaaltiel, Yoseph.
Afiliação
  • Hanania U; Protalix Biotherapeutics, Carmiel, Israel.
  • Ariel T; Protalix Biotherapeutics, Carmiel, Israel.
  • Tekoah Y; Protalix Biotherapeutics, Carmiel, Israel.
  • Fux L; Protalix Biotherapeutics, Carmiel, Israel.
  • Sheva M; Protalix Biotherapeutics, Carmiel, Israel.
  • Gubbay Y; Protalix Biotherapeutics, Carmiel, Israel.
  • Weiss M; Protalix Biotherapeutics, Carmiel, Israel.
  • Oz D; Protalix Biotherapeutics, Carmiel, Israel.
  • Azulay Y; Protalix Biotherapeutics, Carmiel, Israel.
  • Turbovski A; Protalix Biotherapeutics, Carmiel, Israel.
  • Forster Y; Protalix Biotherapeutics, Carmiel, Israel.
  • Shaaltiel Y; Protalix Biotherapeutics, Carmiel, Israel.
Plant Biotechnol J ; 15(9): 1120-1129, 2017 Sep.
Article em En | MEDLINE | ID: mdl-28160363
Plant-produced glycoproteins contain N-linked glycans with plant-specific residues of ß(1,2)-xylose and core α(1,3)-fucose, which do not exist in mammalian-derived proteins. Although our experience with two enzymes that are used for enzyme replacement therapy does not indicate that the plant sugar residues have deleterious effects, we made a conscious decision to eliminate these moieties from plant-expressed proteins. We knocked out the ß(1,2)-xylosyltranferase (XylT) and the α(1,3)-fucosyltransferase (FucT) genes, using CRISPR/Cas9 genome editing, in Nicotiana tabacum L. cv Bright Yellow 2 (BY2) cell suspension. In total, we knocked out 14 loci. The knocked-out lines were stable, viable and exhibited a typical BY2 growing rate. Glycan analysis of the endogenous proteins of these lines exhibited N-linked glycans lacking ß(1,2)-xylose and/or α(1,3)-fucose. The knocked-out lines were further transformed successfully with recombinant DNaseI. The expression level and the activity of the recombinant protein were similar to that of the protein produced in the wild-type BY2 cells. The recombinant DNaseI was shown to be totally free from any xylose and/or fucose residues. The glyco-engineered BY2 lines provide a valuable platform for producing potent biopharmaceutical products. Furthermore, these results demonstrate the power of the CRISPR/Cas9 technology for multiplex gene editing in BY2 cells.
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Texto completo: 1 Base de dados: MEDLINE Assunto principal: Nicotiana / Xilose / Terapia Biológica / Glicoproteínas / Fucose Idioma: En Revista: Plant Biotechnol J Ano de publicação: 2017 Tipo de documento: Article País de afiliação: Israel

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Nicotiana / Xilose / Terapia Biológica / Glicoproteínas / Fucose Idioma: En Revista: Plant Biotechnol J Ano de publicação: 2017 Tipo de documento: Article País de afiliação: Israel