title: Engineering of complex protein sialylation in plants.
abstract: Sialic acids (Sias) are abundant terminal modifications of protein-linked glycans. A unique feature of Sia, compared with other monosaccharides, is the formation of linear homo-polymers, with its most complex form polysialic acid (polySia). Sia and polySia mediate diverse biological functions and have great potential for therapeutic use. However, technological hurdles in producing defined protein sialylation due to the enormous structural diversity render their precise investigation a challenge. Here, we describe a plant-based expression platform that enables the controlled in vivo synthesis of sialylated structures with different interlinkages and degree of polymerization (DP). The approach relies on a combination of stably transformed plants with transient expression modules. By the introduction of multigene vectors carrying the human sialylation pathway into glycosylation-destructed mutants, transgenic plants that sialylate glycoproteins in α2,6- or α2,3-linkage were generated. Moreover, by the transient coexpression of human α2,8-polysialyltransferases, polySia structures with a DP >40 were synthesized in these plants. Importantly, plant-derived polySia are functionally active, as demonstrated by a cell-based cytotoxicity assay and inhibition of microglia activation. This pathway engineering approach enables experimental investigations of defined sialylation and facilitates a rational design of glycan structures with optimized biotechnological functions.
authors.affiliation: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences, 1190 Vienna, Austria;
authors.forename: Somanath
authors.initials: S
authors.lastname: Kallolimath
authors.affiliation: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences, 1190 Vienna, Austria;
authors.forename: Alexandra
authors.initials: A
authors.lastname: Castilho
authors.affiliation: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences, 1190 Vienna, Austria;
authors.forename: Richard
authors.initials: R
authors.lastname: Strasser
authors.affiliation: Department of Chemistry, University of Natural Resources and Life Sciences, 1190 Vienna, Austria;
authors.forename: Clemens
authors.initials: C
authors.lastname: Grünwald-Gruber
authors.affiliation: Department of Chemistry, University of Natural Resources and Life Sciences, 1190 Vienna, Austria;
authors.forename: Friedrich
authors.initials: F
authors.lastname: Altmann
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Giessen, D-35392 Giessen, Germany;
authors.forename: Sebastian
authors.initials: S
authors.lastname: Strubl
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Giessen, D-35392 Giessen, Germany;
authors.forename: Christina Elisabeth
authors.initials: CE
authors.lastname: Galuska
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Giessen, D-35392 Giessen, Germany;
authors.forename: Kristina
authors.initials: K
authors.lastname: Zlatina
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, University of Giessen, D-35392 Giessen, Germany; Institute of Reproductive Biology, Leibniz Institute for Farm Animal Biology, D-18196 Dummerstorf, Germany;
authors.forename: Sebastian Peter
authors.initials: SP
authors.lastname: Galuska
authors.affiliation: Nambawan Biotech GmbH, D-06120 Halle/Saale, Germany;
authors.forename: Stefan
authors.initials: S
authors.lastname: Werner
authors.affiliation: Institute for Cellular Chemistry, Hannover Medical School, 30625 Hannover, Germany.
authors.forename: Hauke
authors.initials: H
authors.lastname: Thiesler
authors.affiliation: Institute for Cellular Chemistry, Hannover Medical School, 30625 Hannover, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Werneburg
authors.affiliation: Institute for Cellular Chemistry, Hannover Medical School, 30625 Hannover, Germany.
authors.forename: Herbert
authors.initials: H
authors.lastname: Hildebrandt
authors.affiliation: Institute for Cellular Chemistry, Hannover Medical School, 30625 Hannover, Germany.
authors.forename: Rita
authors.initials: R
authors.lastname: Gerardy-Schahn
authors.affiliation: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences, 1190 Vienna, Austria; herta.steinkellner@boku.ac.at.
authors.forename: Herta
authors.initials: H
authors.lastname: Steinkellner
keywords: Nicotiana benthamiana; glycoengineering; in planta sialylation; protein polysialylation; recombinant proteins