The reversible macrocyclization of Tyrocidine A aldehyde: a hemiaminal reminiscent of the tetrahedral intermediate of macrolactamization

Abstract

In spite of the important role of peptide macrocyclizations for the generation of conformationally constrained biological ligands, our knowledge of factors that determine the inclination of a substrate to cyclize is low. Therefore, methods that give access to the thermodynamic characterization of these processes are desirable. In this work, we present the isosteric substitution of the amide ligation site of a cyclopeptide by an imine. Applied to the decapeptide antibiotic Tyrocidine A (TycA), the reversible cyclization of the linear aldehyde TycA–CHO resulted in the unexpectedly stable hemiaminal Ψ[CH(OH)NH]–TycA, which is equivalent to the tetrahedral intermediate of macrolactamization, and which is observed for the first time in a peptidic structure. On the basis of NMR spectroscopy and molecular modeling, we discuss the observed high stereoselectivity of hemiaminal formation, as well as its reluctance to be dehydrated to the imine. As required for thermodynamic analysis, it is possible to establish a pH- and temperature-dependent cyclization equilibrium, which allows determination of the entropy loss of the peptide chain, and quantification of the extent of preorientation of the cyclization precursor.