Three in one: Native chemical ligation (NCL) and bis(2‐sulfanylethyl)amido (SEA) ligation allow the one‐pot assembly of three peptide segments in the N‐to‐C direction. The SEA group (see picture, blue) is switched off by intramolecular disulfide bond formation during NCL. Then, a phosphine switches it on to trigger the second SEA ligation step. The K1 domain of the hepatocyte growth factor was synthesized and found to be biologically active.
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.
Drei in einem: Native chemische Ligation (NCL) und Bis(2‐sulfanylethyl)amido(SEA)‐Ligation ermöglichen die Kupplung von drei Peptidsegmenten in einem Gefäß in N‐nach‐C‐Richtung. Die SEA‐Gruppe (siehe Bild, blau) liegt während der NCL als intramolekulares Disulfid vor. Für den zweiten SEA‐Ligationsschritt wird sie in situ mit einem Phosphin angeschaltet. Die K1‐Domäne des Hepatozyten‐Wachstumsfaktors wurde so synthetisiert und erwies sich als biologisch aktiv.
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