N‐Sulfanylethylanilide Peptide as a Crypto‐Thioester Peptide

Abstract

Native chemical ligation (NCL) has shown great utility in protein chemistry and has yielded impressive success in the preparation of a wide variety of proteins.[1] This methodology requires peptide thioesters that serve as chemoselective acylating agents for N-terminal cysteinyl peptides to afford ligated peptides through a sequence of reactions consisting of S-S and S-N acyl transfers. The susceptibility of the thioester moiety to basic reagents has necessitated the preparation of the key intermediate by Boc-based solid-phase peptide synthesis (Boc-SPPS) without requiring a nucleophile-mediated deprotection procedure.[2] However, the preferred use of Fmoc-based SPPS with piperidine treatment demands the development of a synthetic methodology using peptide thioesters that are compatible with Fmoc chemistry.In this context, many research groups, including ours, have explored an Fmoc-based synthetic protocol for thioesters. [3][4][5] Among the reported studies, N-S acyl-transfer-mediated procedures have great potential in Fmoc chemistry.[5] We have also developed an N-sulfanylethylaniline linker that can be used for the acyl-transfer-mediated synthesis of peptide thioesters.[5g]Standard Fmoc-SPPS on the sulfanylethylaniline linker followed by N-S acyl transfer under acidic conditions (4 m HCl in DMF) efficiently yielded peptide thioesters (Scheme 1). On the basis of these experimental results, we attempted to utilize an N-terminal cysteinyl N-sulfanylethylanilide (SEAlide) peptide as the middle fragment(s) for sequential NCL, which features the use of more than one thioester fragment.[6] Here, involvement of the SEAlide peptide in the first NCL with a peptide thioester would seem to selectively afford the corresponding ligated SEAlide peptide, which can be used in the second NCL step after conversion of the anilide moiety to the thioester under acidic conditions. The first NCL doubtlessly proceeded; however, contrary to our expectations, a not insignificant amount of cyclic material resulting from the unanticipated intramolecular NCL of the cysteinyl SEAlide peptide was observed (Scheme 2). This unexpected result indicated that the SEAlide moiety could work as a thioester in the presence of an N-terminal cysteinyl residue even under neutral NCL conditions to afford the corresponding NCL product. [7] In this study, we first examined the feasibility of the SEAlide peptide as a crypto-thioester peptide, and discuss the utility of the cryptic thioester in a kinetically controlled NCL. [8] Initial evaluation of the utility of the SEAlide peptide under NCL conditions was attempted through model coupling reactions between peptide 1 and 2 (Table 1). After preliminary experiments, we first fixed the control coupling conditions (1 mm each peptide in 6 m guanidine·HCl (Gn·HCl)-0.2 m sodium phosphate in the presence of 100 mm (4-carboxymethyl)thiophenol (MPAA) [9,10] and 40 mm tris(2-carboxyethyl)phosphine (TCEP), pH 7.3, 37 8C). Under standard conditions, the attempted NCL between 1 a and 2 was almost complete in 48 h and ...

Efficient one-pot synthesis of CXCL14 and its derivative using an N -sulfanylethylanilide peptide as a peptide thioester equivalent and their biological evaluation

“…23 Previous synthetic strategies involved several experimental manipulations including purification after the first NCL, removal of the N-terminal cysteine protection (C-to-N) and conversion of thioacid to thioester (N-to-C), which resulted in low overall yields. Recently, we discovered that the N-sulfanylethylanilide (SEAlide) peptide efficiently functions as a thioester equivalent in the presence of phosphate salts to participate in NCL [24][25][26] (Scheme 1a). This feature enables the SEAlide peptide to enhance the performance of an N-to-C-directive one-pot/sequential NCL protocol (Scheme 1b).…”

From protein total synthesis to peptide transamidation and metathesis: playing with the reversibility of N,S-acyl or N,Se-acyl migration reactions

“…This reaction involves two sequential rearrangements since the process starts with an N,S-acyl shift of the SEA tertiary amide group, and finishes with an S,N-acyl shift at the level of the transient thioesterlinked intermediate formed by a classical NCL reaction mechanism. Other studies showed that the capacity of SEA peptides to be transamidated by N-terminal cysteinyl peptides in water at a pH close to neutrality is shared by other N,S-acyl shift systems such as the SEAlide [28], the N-alkylcysteine [15,29 ] or the a-methyl cysteine The design of selective peptide transamidation (a) or amide metathesis (b) reactions is highly challenging. Up to now, peptide-based dynamic covalent libraries relied on non-peptidic reversible reactions such as the thiol/disulfide (c), the amine/imine (d) or the thiol/thioester exchange reactions (e).…”

Selectively Activatable Latent Thiol and Selenolesters Simplify the Access to Cyclic or Branched Peptide Scaffolds

“…10,11 Another study relied on the differential reactivity of thiolester surrogates for setting up a kinetically controlled ligation scheme. 9 Fundamentally, the process described in Figure 4 differs from the previous studies in several aspects. First, only one carbonyl functionality can react at a time, i.e., the process is not kinetically controlled.…”