Oral Presentation 6th Modern Solid Phase Peptide Synthesis & Its Applications Symposium 2017

Development and application of thiol-additive-free native chemical ligation techniques combined with several Cys modifications (#8)

Shugo Tsuda 1 , Ken Sakamoto 1 , Masayoshi Mochizuki 1 , Hideki Nishio 1 , Taku Yoshiya 1
  1. Peptide Institute, Inc., Ibaraki, Osaka, Japan

The total chemical synthesis of peptides/proteins by native chemical ligation (NCL) has contributed much to the biological and medicinal fields. In addition, combining NCL with a desulfurization reaction using radical species has increased its synthetic scope. However, aryl thiol additives such as MPAA used for NCL should be removed by HPLC before desulfurization, because aryl thiol compounds are known to terminate radical reactions that are essential for desulfurization. Under these circumstances, in order to achieve one-pot desulfurization or Cys modification after NCL, we have developed several thiol-additive-free NCL techniques. First, we disclosed that Cys-rich peptides can be efficiently ligated even in the absence of any thiol additives [1]. The method was later applied to the synthesis of Ala-rich peptides with intentional introduction of Cys [2]. In this case, we adopted combination of thiol-additive-free NCL/desulfurization. Next, we developed “TfaC-thioester” using side-chain thiol of N-trifluoroacetyl-Cys (TfaC) [3]. Peptide-TfaC-thioester can be directly synthesized by Boc SPPS. Meanwhile, peptide-TfaC-thioester can participate in the thiol-additive-free NCL reaction as a more active species than the ordinary peptide-alkylthioester owing to an appropriate pKa of the thiol moiety. This method can be also applied to kinetically controlled ligation. Finally, we reported that imidazole [4] or triazole [5] can facilitate NCL as efficiently as MPAA. Using imidazole/triazole-aided NCL, one-pot desulfurization or Cys-modifications can be readily achieved. Skillful combination of these methods and other known techniques will help effective protein preparation by NCL.

 [1] S. Tsuda et al. Organic Letters 2015, 17, 1806. [2] S. Tsuda et al. ChemBioChem 2016, 17, 2133. [3] S. Tsuda et al. Biopolymers (Pept. Sci), 2016, 106, 503. [4] K. Sakamoto et al. Chem. Eur. J. 2016, 22, 17940. [5] K. Sakamoto et al. manuscript in preparation.