title: Transient Glyco-Engineering to Produce Recombinant IgA1 with Defined N- and O-Glycans in Plants.
abstract: The production of therapeutic antibodies to combat pathogens and treat diseases, such as cancer is of great interest for the biotechnology industry. The recent development of plant-based expression systems has demonstrated that plants are well-suited for the production of recombinant monoclonal antibodies with defined glycosylation. Compared to immunoglobulin G (IgG), less effort has been undertaken to express immunoglobulin A (IgA), which is the most prevalent antibody class at mucosal sites and a promising candidate for novel recombinant biopharmaceuticals with enhanced anti-tumor activity. Here, we transiently expressed recombinant human IgA1 against the VP8* rotavirus antigen in glyco-engineered ΔXT/FT Nicotiana benthamiana plants. Mass spectrometric analysis of IgA1 glycopeptides revealed the presence of complex biantennary N-glycans with terminal N-acetylglucosamine present on the N-glycosylation site of the CH2 domain in the IgA1 alpha chain. Analysis of the peptide carrying nine potential O-glycosylation sites in the IgA1 alpha chain hinge region showed the presence of plant-specific modifications including hydroxyproline formation and the attachment of pentoses. By co-expression of enzymes required for initiation and elongation of human O-glycosylation it was possible to generate disialylated mucin-type core 1 O-glycans on plant-produced IgA1. Our data demonstrate that ΔXT/FT N. benthamiana plants can be engineered toward the production of recombinant IgA1 with defined human-type N- and O-linked glycans.
authors: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences Vienna, Austria.
authors: Martina
authors: M
authors: Dicker
authors: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences Vienna, Austria.
authors: Marc
authors: M
authors: Tschofen
authors: Department of Chemistry, University of Natural Resources and Life Sciences Vienna, Austria.
authors: Daniel
authors: D
authors: Maresch
authors: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences Vienna, Austria.
authors: Julia
authors: J
authors: König
authors: Institute of Molecular and Cellular Plant Biology, Spanish Research Council Agency - Polytechnic University of Valencia Valencia, Spain.
authors: Paloma
authors: P
authors: Juarez
authors: Institute of Molecular and Cellular Plant Biology, Spanish Research Council Agency - Polytechnic University of Valencia Valencia, Spain.
authors: Diego
authors: D
authors: Orzaez
authors: Department of Chemistry, University of Natural Resources and Life Sciences Vienna, Austria.
authors: Friedrich
authors: F
authors: Altmann
authors: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences Vienna, Austria.
authors: Herta
authors: H
authors: Steinkellner
authors: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences Vienna, Austria.
authors: Richard
authors: R
authors: Strasser
keywords: N-glycosylation; O-glycosylation; antibody; glyco-engineering; monomeric IgA; plant-made pharmaceuticals; protein glycosylation; recombinant glycoprotein