In a modern chemical protein synthesis, the polypeptide backbone of target protein is assembled by chemoselective ligation of peptide segments. In principle, these reactions require leaving group at the C-terminus of one peptide to couple with another peptide segment. So far, peptide-^αthioester has been utilized as the C-terminal activated derivative. Peptide-^αthioester have played a pivotal role in chemical protein synthesis, however, a less choice of the leaving group would limit the synthetic strategy. For example, it could be still troublesome to perform kinetically controlled ligation (KCL) between peptide-^αthioesters unless suppressing the reactivity of one thioester or choosing ligation site carefully. Thus, we set out to develop a new leaving group that is compatible with thioester but can exhibit different reactivity to expand the orthogonal peptide assemble strategy for chemical protein synthesis.

In such a context, we have found a unique peptide derivative: peptidyl-N-acylguanidine, which has N-acylguanidine group at the C-terminus of peptide and exhibits much less reactivity than that of thioester in KCL^[1]. Intriguingly, peptidyl-N-acylguanidine could be converted into peptide-^αthioester in the neutral buffer solutions containing excess amount of alkyl thiol. This unique reactivity enabled us to perform an efficient KCL between peptide-^αthioester and peptidyl-N-acylguanidine and following trans-thioesterification of N-acylguanidine moiety. We have also successfully developed the efficient synthetic methods of peptidyl-N-acylguanidine based on conventional Fmoc and Boc solid phase peptide synthesis (SPPS). The unique property of peptidyl-N-acylguanidine and related chemistry may open a new way for the chemical protein synthesis and enable us to synthesize further complicated protein structures.

[1] R. Okamoto, K. Morooka, Y. Kajihara, Angew. Chem. Int. Ed. 2012, 51, 191-197