FULL PAPER DOI: 10.1002/ejoc.201300050 All-Thioamidated Homo-α-Peptides: Synthesis and Conformation Fernando Formaggio,* [a] Marco Crisma, [a] Claudio Toniolo, [a] and Cristina Peggion* [a] Keywords: Conformation analysis / Synthetic methods / Peptides / Peptidomimetics Replacement of a peptide bond with its thioamide surrogate is a classical method for the generation of a peptidomimetic with altered spectroscopic, conformational, physicochemical, and biological properties. In this context, we synthesized short series of terminally protected homo-α-oligopeptides based on the α-amino acids Gly, Ala, and Nle, as well as their corresponding fully thioamidated analogues. For the first Introduction Thionated peptides are members of an interesting class of backbone-modified peptidomimetics [1] that have been utilized in a variety of fields, including medicinal chemistry, spectroscopy, and photophysics (cis/trans thioamide isomer- ization). [1–25] Their preferred conformations have been ex- tensively investigated, in particular by X-ray diffrac- tion, NMR, and electronic and energy calcula- tions. [2,3,7,13,15,17,26–59] Most of these studies have involved compounds with single, site-specific peptide bond modifica- tions. These targets have typically been achieved by regiose- lective thionation of very short peptides (e.g., terminally protected tripeptides), followed by extension of the amino acid chain by chemical synthesis. [2,3,30,60–65] The classical protecting functionalities used in peptide synthesis (ureth- anes at N termini and esters at C termini) are not modified under the experimental conditions almost universally em- ployed for thionation of an amide group (Lawesson rea- gent [60] in an organic solvent of low polarity). Moreover, additional regioselectivity between two (or three) amide groups can frequently be achieved by taking advantage of the differences in steric hindrance exerted by the side chains of the pairs of surrounding amino acids. In contrast, only a very limited number of articles deal- ing with synthesis and conformation of all-thioamidated peptides have appeared in the literature. [2,28] This observa- tion is rather surprising in view of the great potential inter- est of this class of peptidomimetics as new foldameric struc- [a] ICB, Padova Unit, CNR, Department of Chemistry, University of Padova, 35131 Padova, Italy E-mail: fernando.formaggio@unipd.it cristina.peggion@unipd.it Homepage: http://www.chimica.unipd.it/toniologroup/ index_file/Page1089.htm Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/ejoc.201300050. Eur. J. Org. Chem. 2013, 3455–3463 © 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim 3455 time, the preparation of the latter compounds was achieved in single-step fashion through direct thionation of their oxy- genated precursors. Using X-ray diffraction analysis and NMR spectroscopy we were also able to confirm that the thioamidated α-amino acid residues can easily adopt either folded or fully extended conformations. tures and the increasing number of naturally occurring poly-α- and -β-peptides containing up to five consecutive thioamide groups in their amino acid chains, that have been isolated and sequenced. [66–68] In this work we decided to fill this gap at least partially by synthesizing a large set of all- thioamidated, selected peptide foldamers, by use of the single-step direct thionation methodology, and by investi- gating their conformational preferences by detailed X-ray diffraction and 2D-NMR analyses. Results and Discussion Peptide Synthesis and Characterization We used classical solution-phase methods to prepare three short series (from dimer through tetramer) of ter- minally protected homo-α-oligopeptides based on the Gly, Ala, and Nle (norleucine) residues. These amino acids differ in their linear, but increasingly bulkier, side chains (R), where R = H for Gly, R = –CH 3 for Ala, and R = –CH 2 – (CH 2 ) 2 –CH 3 for Nle. The syntheses and characterizations of these nine peptides had already been reported. [69–72] Thionation of the Gly dipeptide with Lawesson reagent (LR) [60] under typical conditions [anhydrous THF at room temperature; Procedure A (see the Exp. Section)] proceeded fairly smoothly in about 1 hour to afford the monothioami- dated compound 2a (Scheme 1) after chromatographic pu- rification (no special efforts were made to optimize the yields in our thionation reactions). To obtain the desired Gly bis-thioamidated product 3a and the tris-thioamidated product 4a, a further aliquot of LR (to a total of 1.8–3.0 equiv.) was added after 24 hours in each case. The reaction mixtures were left whilst stirring for additional 24 hours. The preparations of the Ala bis-thioamidated tripeptide 3b and the tris-thioamidated tetrapeptide 4b (but not that