Serine-, threonine-, and cysteine-derived cyclic building blocks (pseudo-prolines, ΨPro) serve as reversible protecting groups for Ser, Thr, and Cys and prove to be versatile tools for overcoming some intrinsic problems in the field of peptide chemistry. The presence of ΨPro within a peptide sequence results in the disruption of β-sheet structures considered as a source of intermolecular aggregation during chain elongation, thus increasing solvation and coupling kinetics in peptide assembly. Due to their easy synthetic access and variability in the chemical stability by modifications introduced in the C-2 position of the oxazolidine/thiazolidine ring system, this protection technique is adaptable to all common strategies in peptide synthesis. We describe new types of ΨPro building blocks suitable for standard Fmoc/tBu-based solid phase peptide synthesis, convergent strategies, and chemoselective ligation techniques as well as their use as a structure-disrupting, solubilizing protection technique for the example of peptides generally considered as “difficult sequences”.

Serine, threonine-derived (4S)-oxazolidine-4-carboxylic acid, and cysteine-derived (4R)-thiazolidinecarboxylic acid, denoted pseudo-proline (Xaa[ΨR 1 ,R 2 pro]), serve as structure disrupting, solubilizing building blocks in peptide synthesis. Variation of the 2-C substituents within the heterocyclic system results in different physicochemical and conformational properties. NMR studies of a series of pseudo-proline (ΨPro)-containing peptides reveal a pronounced effect of the 2-C substituents upon the cis to trans ratio of the adjacent amide bond in solution. 2-C unsubstituted systems show a preference similar to that of the proline residue for the trans form, whereas 2,2-dimethylated derivatives adopt the cis amide conformation in high content. For 2-monosubstituted ΨPro, the cis−trans distribution depends on the 2-C chirality. For the 2-(S)-diastereoisomer, both forms are similarly populated in solution, whereas the 2-(R)-epimer adopts preferentially the trans form. The results are supported by conformational energy calculations and suggest that, by tailoring the degree of substitution, pseudo-prolines may serve as a temporary proline mimetic or as a hinge in peptide backbones.

Molecular events such as cis/trans isomerization of Xaa‐Pro tertiary amide bonds in peptides and proteins are slow on the overall time scale of the formation of a final biostructure and are, therefore, rate limiting. In order to pursue a better understanding of the molecular events underlying such slow interconversions, we applied the recently introduced pseudo‐proline (ΨPro) concept as a tool to study the dynamics of Xaa‐Pro bonds by determining the kinetics and thermodynamics of cis/trans isomerism. We show that enhanced isomerization rates of tertiary amide bonds prior to a ΨPro unit in short model peptides is due to lowered transition state barriers. In addition, pronounced effects upon the dynamics of the reversible transition between helix I and II of oligoprolines containing one or several ΨPro units were observed. © 1999 John Wiley & Sons, Inc. Biopoly 51: 121–128, 1999

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The insertion of acetals that exhibit variable structural features into complex peptides such as cyclosporin C (CsC) results in oxazolidine derivatives (pseudoprolines, psiPro) of tailored physico-chemical and biological properties. N,O-Acetalation of the 2-threonine hydroxyl group and the preceding amide nitrogen of CsC is achieved by treating the molecule with a number of both arylated and non-arylated dimethyl acetals. The psiPro-containing CsC derivatives exhibit enhanced conformational backbone rigidity, as suggested by analytical HPLC, NMR spectroscopy and by kinetic measurements on binding with their receptor protein cyclophilin A (CypA) that were not time-dependent. IC50 values for calf-thymus CypA were obtained by kinetic evaluation of its cis-->trans isomerase activity. The choice of the para-substituted aryl dimethyl acetals allows the inhibitory properties of the corresponding derivatives to be modulated to either prodrugs or moderately strongly binding cyclosporin C derivatives.