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Evaluation of Immunoglobulins from Plant Cells
Author(s) -
Hein Mich B,
Tang Ying,
Mcleod Donald A,
Janda Kim D,
Hiatt Andrew
Publication year - 1991
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp00011a011
Subject(s) - endoglycosidase , endoglycosidase h , glycan , antibody , biochemistry , endomembrane system , biology , glycoprotein , lectin , microbiology and biotechnology , monoclonal antibody , golgi apparatus , antigen , complementary dna , gene , endoplasmic reticulum , genetics , immunology
Expression of cDNA constructs encoding full‐length mouse immunoglobulin chains with their native leader sequences or fusion constructs substituting the native leader with a pre‐pro sequence derived from Saccharomyces cerevisiae yielded blocked N‐termini on the γ chain or the correct amino terminal sequence on the mature k chain. Lectin binding assays revealed that assembled immunoglobulin complexes contained a glycosylated heavy chain. The attached glycan was resistant to digestion by endoglycosidase H and its lectin binding pattern was distinguishable from that of the mammalian glycan. The results indicated processing of the immunoglobulin carbohydrate in the tobacco Golgi to yield a complex oligosaccharide. Secretion of antibody by protoplasts isolated from regenerated transgenic plants or from suspension callus cells was demonstrated by pulse‐chase labeling experiments. When purified, the tobacco‐produced antibody was found to possess the antigen binding and catalytic properties of the murine monoclonal antibody. Kinetic parameters ( K m , K i , V max , and k cat ) of the tobacco‐derived antibody were comparable to those of the mouse‐derived antibody. The results in general show that the endomembrane system of tobacco cells possesses cognate mechanisms for the recognition of diverse leader sequences. These signals can be used to initiate the assembly, processing, and secretion by plant cells of complex foreign proteins.