Protease-mediated peptide bond formation. On some unexpected outcomes during enzymatic synthesis of leu-enkephalin.
In the course of a study in protease-controlled peptide synthesis, several promising pathways to synthetic enkephalins had to be discarded or modified because they failed to give the required products. Partial deprotection of peptide fragments prior to chain elongation resulted in an enhanced susceptibility of scissile bonds to proteolysis. The use of proteases, the specificity of which was not confined solely to the bond to be synthesized, thus led to a priori cleavage of pre-existing peptide bonds. Subsequently, enzyme-mediated synthesis of new peptide bonds between initial reactants and nascent degradation products furnished undesired, often truncated peptides. A detailed characterization of the undesired products gave information as to whether or not the peptide bonds were susceptible to proteolytic cleavage or accessible via enzymatic synthesis. In this way, the unexpected outcome of protease catalysis led to working predictions regarding enzyme-mediated peptide bond formation, and thus, finally contributed to the successful synthesis of the target peptides.