• Open Access

A KDEL-tagged monoclonal antibody is efficiently retained in the endoplasmic reticulum in leaves, but is both partially secreted and sorted to protein storage vacuoles in seeds

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* Correspondence (fax 54-221-425-4854; e-mail silvana@biol.unlp.edu.ar)

Summary

Transgenic plants are attractive biological systems for the large-scale production of pharmaceutical proteins. In particular, seeds offer special advantages, such as ease of handling and long-term stable storage. Nevertheless, most of the studies of the expression of antibodies in plants have been performed in leaves. We report the expression of a secreted (sec-Ab) or KDEL-tagged (Ab-KDEL) mutant of the 14D9 monoclonal antibody in transgenic tobacco leaves and seeds. Although the KDEL sequence has little effect on the accumulation of the antibody in leaves, it leads to a higher antibody yield in seeds. sec-AbLeaf purified from leaf contains complex N-glycans, including Lewisa epitopes, as typically found in extracellular glycoproteins. In contrast, Ab-KDELLeaf bears only high-mannose-type oligosaccharides (mostly Man 7 and 8) consistent with an efficient endoplasmic reticulum (ER) retention/cis-Golgi retrieval of the antibody. sec-Ab and Ab-KDEL γ chains purified from seeds are cleaved by proteases and contain complex N-glycans indicating maturation in the late Golgi compartments. Consistent with glycosylation of the protein, Ab-KDELSeed was partially secreted and sorted to protein storage vacuoles (PSVs) in seeds and not found in the ER. This dual targeting may be due to KDEL-mediated targeting to the PSV and to a partial saturation of the vacuolar sorting machinery. Taken together, our results reveal important differences in the ER retention and vacuolar sorting machinery between leaves and seeds. In addition, we demonstrate that a plant-made antibody with triantennary high-mannose-type N-glycans has similar Fab functionality to its counterpart with biantennary complex N-glycans, but the former antibody interacts with protein A in a stronger manner and is more immunogenic than the latter. Such differences could be related to a variable immunoglobulin G (IgG)-Fc folding that would depend on the size of the N-glycan.

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