Applying our thioesterification methods, the Okamoto and Kajihara group synthesized interleukin 13 (IL-13) in 2014 (Fig15). [42] At the first they reported a new semi-synthetic strategy of IL-13 in which the middle region is glycosylated.46 IL-13 is a glycoprotein that consists of 112 amino acids and has a complex-type oligosaccharide at Asn52. According to their synthetic strategy of IL-13, they employed four peptide segments and three sequential NCL to obtain a full-length glycoprotein. Then IL-13 segment 133 (1-27) and 136 (55-112) were simultaneously prepared by E. coli. expression system and cleaved fusion proteins 131 by CNBr, which can selectively cleave an amide bond between the Met and Cys residues; furthermore, the Met residue is converted into the homoserine lactone (Hsl) (Fig.15). They utilized cleavage reaction on the amide bond between artificially inserted Met and Cys with CNBr to afford segment 133 (1-27) and 136 (55-112) simultaneously in high yield. Then segment 133 (1-27) was converted to peptide thioesters 138 by using their developed “guanidine method” (Fig.14, A) and obtained peptide α thioester. On the other hands, glycopeptide thioester 135 was synthesized by Fmoc standard SPPS protocol followed by their reported manner. After preparation of all segments by semisynthetic approaches, sequential NCL was convergently progressed to afford the linear full-length IL13 segment139. Folding process under stepwise dialysis with redox reagents yielded folded IL13 140. ELISA and CD spectrum indicated that semi-synthesized IL13 correctly folded having α-helix rich conformation.

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