Enzymatic synthesis of X-Phe-Leu-NH₂ in low water content systems: Influence of the N-α protecting group and the reaction medium composition

The influence of eight different N-terminal protecting groups (For, Ac, Boc, Fmoc, Mal, Pheac, Aloc and Z) on the α-chymotrypsin-catalyzed synthesis of the dipeptide derivative X-Phe-Leu-NH₂ in organic media was studied. Groups such as Ac, For, Boc, Z, Mal, Pheac and Alloc always rendered good peptide yields (92% to 99%) either in acetonitrile or in ethyl acetate. Good correlations were found between molecular and physico-chemical characteristics of the N-α moiety such as the hydrophobicity (log P), ovality and dipole moment and the global reaction rate parameter k′. High k′ values were obtained with the less hydrophobic groups, Ac, For and Mal, that have ovality values close to one and the highest dipole moments. Furthermore, it was found that the relative rate of hydrolysis and aminolysis of the acyl-enzyme intermediate expressed as the partition parameter p is affected by the N-α moiety of the acyl donor. Correlations between this parameter and the dipole moment of the protecting group were observed.