Enzymatic Semisynthesis of Dicarba Analogs of Calcitonin

The semisynthesis of eel[L‐α‐aminosuberic acid1.7]calcitonin (elcatonin) was accomplished by α‐chymotrypsin‐catalyzed coupling of two peptide segments in a single reaction without the protection of any functional group. The eel calcitonin‐(10–32)‐peptide was prepared by a gene manipulation. The esters of cyclic desamino nonapeptide (segment 1–9) were synthesized by the conventional solution method including a thermolysin‐mediated resolution of DL‐α‐aminosuberic acid via one‐step tripeptide synthesis leading to the 7–9 sequence. The main aim of this work was to determine the conditions for protease‐catalyzed segment condensation while avoiding a concurrent cleavage of other proteolytically labile peptide bonds in the hormone. The α‐chymotrypsin condensation strategy under usual conditions led to a complicated mixture of split products with an insignificant amount of the required peptide. When the coupling reaction was carried out at 0°C, the reaction resulted in a satisfactory yield of elcatonin with the complete conversion of the acyl donor (1–9 segment) accompanied by negligible concurrent peptide bond digestion. The same strategy was employed for the preparation of analogous dicarba salmon calcitonin using a synthetic elcatonin‐(10–32)‐peptide. Both calcitonin analogs exhibited hypocalcemic activity corresponding to the international standard of elcatonin. We demonstrate in this work a peptide synthesis based on the combination of genetic engineering, chemical synthesis and proteinase‐catalyzed segment condensation. This approach enables effective incorporation of an unnatural amino acid into calcitonins without the side‐chain protection.