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N‐linked glycosylation of IL‐13R0'2 is essential for optimal IL‐13 inhibitory activity
Author(s) -
Kioi Mitomu,
Seetharam Saraswathy,
Puri Raj K.
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.06-5995fje
Subject(s) - glycosylation , microbiology and biotechnology , biochemistry , chemistry , biological activity , receptor , cell culture , biology , in vitro , genetics
A high‐affinity receptor for interleukin (IL)‐13 (interleukin‐13Rα 2) is over‐expressed in disease‐related fibroblasts and neoplastic cells and is involved in cancer, allergic, and inflammatory diseases. The extracellular domain of IL‐13Rα2 (ECDα2) could be cleaved, which serves as a decoy receptor. We have expressed and purified ECDα2 in both Escherichia coli ( E. coli ) and mammalian systems as a soluble fragment and studied its biological activities. Although both products of ECDα2 showed IL‐13 inhibitory activities, mammalian cell‐derived ECDα2 appeared to be superior compared with purified protein from E. coli . When expressed in E. coli , ECDα2 appeared to be a monomer of 42 but a 60 kDa protein when purified from mammalian cells due to heavy glycosylation. The purified glycosylated ECDα2 efficiently inhibited IL‐13‐induced STAT6 phosphorylation in immune and Hodgkin's lymphoma cell lines, IL‐13 binding, and cytotoxicity of IL‐13 cytotoxin in various cancer cell lines. The improved potency of mammalian cell‐derived ECDα2 was shown over ECDα2/Fc fusion protein. The N ‐linked glycosylation of ECDα2 was found to be essential for optimal IL‐13 inhibitory activity as deglycosylation by PNGase F showed lower activity. ECDα2 did not inhibit IL‐4‐induced STAT6 phosphorylation, indicating that inhibitory effects of ECDα2 are receptor specific. These results indicate that glycosylated ECDα2 can serve as a potent inhibitor of IL‐13 in a variety of conditions in which IL‐13 is a key mediator, e.g ., pulmonary, allergic, fibrotic, and neoplastic diseases.—Kioi, M., Seetharam, S., Puri, R. K. N‐linked glycosylation of IL‐13Rα2 is essential for optimal IL‐13 inhibitory activity. FASEB J. 20, E1670 –E1680 (2006)