Semi-synthesis of homogeneous glycoproteins via novel expressed peptide thioesterification methods — ASN Events

Semi-synthesis of homogeneous glycoproteins via novel expressed peptide thioesterification methods (#12)

Yasuhiro Kajihara 1
  1. Department of Chemistry, Osaka University, Machikaneyama, Toyonaka, Japan

The elucidation of glycosylation function usually requires homogeneous glycoproteins with structurally defined glycans. However, currently, the mammalian cell expression only produce glycoprotein with heterogeneous glycan structure. On the other hand, total chemical synthesis of glycoprotein based on solid-phase peptide synthesis (SPPS) is time-consuming. Semi-synthesis combining E. coli expression and chemical synthesis appears to be a powerful approach to obtain homogenous glycoprotein.

The thioesterification of expressed peptide is an essential step for glycoprotein semi-synthesis. Recently, we have developed several chemical methods for expressed peptide thioesterification based on activation of additional C-terminal Cys. In the first method, the C-terminal Cys was activated through N-S acyl shift and converted to bis(2-sulfanylethyl)amide (SEA) to yield peptide thioester surrogate. The semi-synthesis of glycoprotein inducible T cell costimulator (ICOS) was succeeded by utilizing the first method1. In the second method, the activation of Cys was carried out by S-cyanylation and hydrazinolysis to obtain peptide hydrazide2,3. Although the presence of internal Cys might hamper this reaction due to lack of regioselectivity, in this research, we achieved regioselective C-terminal activation via a novel method “folding assisted thioesterification”. This method specifically accelerated the formation of native disulfide bonds to make a free cysteine at the C-terminal. Toward this Cys residues, hydrazinolysis was successfully performed. Based on this method, we succeeded in semi-synthesis of N143-glycosylated interleukin-6 (IL-6). We suppose that these novel expressed peptide thioesterification methods could greatly facilitate the semi-synthesis of homogeneous glycoproteins, especially those challenging synthetic targets.

  1. R. Okamoto, et al., J. Org. Chem. 2022, 87, 114-124
  2. Y.K. Chong et al., Org. Biomol. Chem., 2022, 20, 1907
  3. Y.B. Liu et al., Current Opinion in Chemical Biology, 2023, 73, 102263
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