Chemical ligation of unprotected peptides has enabled the successful synthesis of a large number of diverse proteins. [1] The native chemical ligation (NCL) reaction, [2] which proceeds in water at neutral pH to yield a native amide bond at the ligation site, has proven to be the most effective chemistry for the ligation of unprotected peptides. A major limitation of protein synthesis by NCL, and indeed all peptide chemicalligation methods, is the steady decrease in yield because of losses from intermediate handling and purification. Although recent one-pot schemes [3] for chemical protein synthesis have helped eliminate some intermediate purification steps and, therefore, increased yields, one-pot reactions can, over time and multiple ligations, lead to a large accumulation of byproducts that are difficult to purify from the target polypeptide. For certain classes of proteins, such as integral membrane proteins, an additional major problem is the poor solubility of many intermediate polypeptide products in the solvents used for ligation. There is currently no effective and general strategy for solubilizing poorly soluble intermediate polypeptides in ligation buffers without introducing modifications to the peptide building blocks. We aimed to perform protein synthesis by NCL on a cross-linked polymer support to address these limitations.
Polymer-supported synthetic chemistry, originally introduced by Merrifield for peptide synthesis, [4] is known to have a number of advantages over traditional solution-phase chemistry. These advantages include simple purification through