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Semisynthesis of Biologically Active Glycoforms of the Human Cytokine Interleukin 6
Author(s) -
Reif Andreas,
Siebenhaar Sebastian,
Tröster Andrea,
Schmälzlein Marina,
Lechner Carolin,
Velisetty Phanindra,
Gottwald Karen,
Pöhner Claudia,
Boos Irene,
Schubert Volker,
RoseJohn Stefan,
Unverzagt Carlo
Publication year - 2014
Publication title -
angewandte chemie
Language(s) - English
Resource type - Journals
eISSN - 1521-3757
pISSN - 0044-8249
DOI - 10.1002/ange.201407160
Subject(s) - semisynthesis , native chemical ligation , glycopeptide , chemistry , peptide , chemical ligation , glycoprotein , glycosylation , ligation , biochemistry , stereochemistry , combinatorial chemistry , chemical synthesis , in vitro , microbiology and biotechnology , biology , antibiotics
Human interleukin 6 (IL‐6) is a potent cytokine with immunomodulatory properties. As the influence of N‐glycosylation on the in vivo activities of IL‐6 could not be elucidated so far, a semisynthesis of homogeneous glycoforms of IL‐6 was established by sequential native chemical ligation. The four cysteines of IL‐6 are convenient for ligations and require only the short synthetic glycopeptide 43–48. The Cys‐peptide 49–183 could be obtained recombinantly by cleavage of a SUMO tag. The fragment 1–42 was accessible by the simultaneous cleavage of two inteins, leading to the 1–42 thioester with the native N‐terminus. Ligation and refolding studies showed that the inherently labile AspPro bond 139–140 was detrimental for the sequential C‐ to N‐terminal ligation. A reversed ligation sequence using glycopeptide hydrazides gave full‐length IL‐6 glycoproteins, which showed full bioactivity after efficient refolding and purification.