TRYPSIN-MEDIATED SEMISYNTHESIS OF SALMON-CALCITONIN

Salmon calcitonin, CT(1-32). NH2, was synthesised by the trypsin-media ted coupling of the peptide fragments CT(1-24) and CT(25-32). NH2, pre pared by conventional Fmoc solid-phase chemistry. Optimal conditions r egarding reaction time course, pH, proportion of catalyst, substrate c oncentration and composition of the reaction medium were determined fr om initial studies on the coupling of CT(1-11) to CT(12-24) and of CT( 12-24) to CT(25-32). NH2. For the final successful semisynthesis, we f ound that it was unnecessary to protect lysine residues not involved i n the coupling, and that secondary hydrolysis at these sites could be prevented by increasing the pH of the reaction medium. The reaction ac hieved equilibrium after 30-45 min, with overall conversion of around 30% of the initial amount of CT(1-24) substrate into product. Yields w ere depressed due to cyclisation of the CT(1-24) substrate via air-oxi dation of the Cys(1) and Cys(7) residues. (C) Munksgaard 1995.