<i>N</i>‑Sulfanylethyl­aminooxy­butyramide (SEAoxy): A Crypto-Thioester Compatible with Fmoc Solid-Phase Peptide Synthesis

An <i>N</i>-sulfanylethyl­amino­oxy­butyramide (SEAoxy) has been developed as a novel thioester equivalent for native chemical ligation. SEAoxy peptide was straightforwardly synthesized by conventional Fmoc solid-phase peptide synthesis without a problem. Moreover, SEAoxy peptide could be directly applied to native chemical ligation owing to the intramolecular <i>N</i>-to-<i>S</i> acyl shift that releases the peptide-thioester <i>in situ</i>. This methodology was successfully applied to the synthesis of two bioactive peptides

Synthetic Procedure for <i>N</i>‑Fmoc Amino Acyl‑<i>N</i>‑Sulfanylethylaniline Linker as Crypto-Peptide Thioester Precursor with Application to Native Chemical Ligation

<i>N</i>-Sulfanylethylanilide (SEAlide) peptides <b>1</b>, obtainable using Fmoc-based solid-phase peptide synthesis (Fmoc SPPS), function as crypto-thioesters in native chemical ligation (NCL), yielding a wide variety of peptides/proteins. Their acylating potential with N-terminal cysteinyl peptides <b>2</b> can be tuned by the presence or absence of phosphate salts, leading to one-pot/multifragment ligation, operating under kinetically controlled conditions. SEAlide peptides have already been shown to be promising for use in protein synthesis; however, a widely applicable method for the synthesis of <i>N</i>-Fmoc amino acyl-<i>N</i>-sulfanylethylaniline linkers <b>4</b>, required for the preparation of SEAlide peptides, is unavailable. The present study addresses the development of efficient condensation protocols of 20 naturally occurring amino acid derivatives to the <i>N</i>-sulfanylethylaniline linker <b>5</b>. <i>N</i>-Fmoc amino acyl aniline linkers <b>4</b> of practical use in NCL chemistry, except in the case of the proline- or aspartic acid-containing linker, were successfully synthesized by coupling of POCl₃- or SOCl₂-activated Fmoc amino acid derivatives with sodium anilide species <b>6</b>, without accompanying racemization and loss of side-chain protection. Furthermore, SEAlide peptides <b>7</b> possessing various C-terminal amino acids (Gly, His, Phe, Ala, Asn, Ser, Glu, and Val) were shown to be of practical use in NCL chemistry