Tidbits for the synthesis of bis(2-sulfanylethyl)amido (SEA) polystyrene resin, SEA peptides and peptide thioesters.

Protein total chemical synthesis enables the atom-by-atom control of the protein structure and therefore has a great potential for studying protein function. Native chemical ligation of C-terminal peptide thioesters with N-terminal cysteinyl peptides and related methodologies are central to the field of protein total synthesis. Consequently, methods enabling the facile synthesis of peptide thioesters using Fmoc-SPPS are of great value. Herein, we provide a detailed protocol for the preparation of bis(2-sulfanylethyl)amino polystyrene resin as a starting point for the synthesis of C-terminal bis(2-sulfanylethyl)amido peptides and of peptide thioesters derived from 3-mercaptopropionic acid.

Access to cyclic or branched peptides using bis(2-sulfanylethyl)amido side-chain derivatives of Asp and Glu.

Bis(2-sulfanylethyl)amido (SEA) side-chain derivatives of aspartic and glutamic acids enable the synthesis of tail-to-side chain cyclic or branched peptides using standard Fmoc-SPPS followed by SEA native peptide ligation.

Polysaccharide microarrays: application to the identification of heparan sulphate mimetics.

The interaction of polysaccharides with proteins modulates or triggers many biological effects. In particular, heparan sulphate proteoglycans (HSPGs) have multiple regulatory interactions with growth factors, enzymes, enzyme inhibitors, and some components of the extracellular matrix. The important role played by HSPGs has motivated the synthesis and selection of HSPG mimetics for modulating the biological activity of HS-binding proteins. We present hereinafter an efficient polysaccharide microarray method that allows the screening of HS-mimetic libraries towards HS-binding growth factors, a major class of polypeptides whose inhibition or potentiation is of high medical interest.

Synthesis of peptide alkylthioesters using the intramolecular N,S-acyl shift properties of bis(2-sulfanylethyl)amido peptides.

The design of novel methods giving access to peptide alkylthioesters, the key building blocks for protein synthesis using Native Chemical Ligation, is an important area of research. Bis(2-sulfanylethyl)amido peptides (SEA peptides) 1 equilibrate in aqueous solution with S-2-(2-mercaptoethylamino)ethyl thioester peptides 2 through an N,S-acyl shift mechanism. HPLC was used to study the rate of equilibration for different C-terminal amino acids and the position of equilibrium as a function of pH. We show also that thioester form 2 can participate efficiently in a thiol-thioester exchange reaction with 5% aqueous 3-mercaptopropionic acid. The highest reaction rate was obtained at pH 4. These experimental conditions are significantly less acidic than those reported in the past for related systems. The method was validated with the synthesis of a 24-mer peptide thioester. Consequently, SEA peptides 1 constitute a powerful platform for access to native chemical ligation methodologies.

Synthesis of thiazolidine thioester peptides and acceleration of native chemical ligation.

Thiazolidine thioester peptides were synthesized by reacting bis(2-sulfanylethyl)amido peptides with glyoxylic acid at pH 1. A significant increase in Native Chemical Ligation (NCL) rate was observed with thiazolidine thioesters compared to 3-mercaptopropionic acid-thioester analogues. The method is of particular interest for accelerating valine-cysteine peptide bond formation.

Bis(2-sulfanylethyl)amino native peptide ligation.

The reaction of a peptide featuring a bis(2-sulfanylethyl)amino (SEA) group on its C-terminus with a cysteinyl peptide in water at pH 7 and 37 °C leads to the chemoselective and regioselective formation of a native peptide bond. This method called SEA ligation enriches the native peptide ligation repertoire available to the peptide chemist. Preparation of an innovative solid support which allows the straightforward synthesis of peptide SEA fragments using standard Fmoc/tert-butyl solid phase peptide synthesis procedures is also described.