SEA Ligation Is Accelerated at Mildly Acidic pH. Application to the Formation of Difficult Peptide Junctions

The bis(2-sulfanylethyl)amido (SEA)-mediated ligation has been introduced in 2010 as a novel chemoselective peptide bond forming reaction. SEA ligation is a useful reaction for protein total synthesis that is complementary to the native chemical ligation (NCL). In particular, SEA ligation proceeds efficiently in a wide range of pH, from neutral pH to pH 3-4. Thus, the pH can be chosen to optimize the solubility of the peptide segments or final product. It can be also chosen to facilitate the formation of difficult junctions, since the rate of SEA ligation increases significantly by decreasing the pH from 7.2 to 4.0. Here we describe a protocol for SEA ligation at pH 5.5 in the presence of 4-mercaptophenylacetic acid (MPAA) or at pH 4.0 in the presence of a newly developed diselenol catalyst. The protocols describe the formation of a valyl-cysteinyl peptide bond between two model peptides.