H j r

brings both aminopyrazole DAD sites roughly in one line (Figure 2.4.6). In 13 two intramolecular hydrogen bonds between the pyridine N-atom and the amide NH-groups in their vicinity fix both aminopyrazole nuclei in a kinked orientation. Although the other ligands can also adopt flat geometries with both heterocyclic moieties pointing in one direction, their distance seems not well suited to interaction with model peptide 14. Especially in the terephthaloyl-bridged dimer the second aminopyrazole cannot be fitted on to the ADA pattern of the peptide. It is, nevertheless, remarkable that dimeric aminopyrazoles are capable of forming multiple hydrogen bonds even with a tetrapeptide containing only alanines, which are known for their relatively low ¿-sheet propensity. In both successful examples more than four hydrogen bonds are formed simultaneously, leading to superior interactions compared with all other aminopyrazole dimers.

The dimeric aminopyrazoles have also been tested on the Prion and Alzheimer's proteins (for details, see Section 2.4.5). Preliminary experiments showed at an early stage of the investigation that even covalent aminopyrazole dimerization produced peptide ligands which could significantly retard protein aggregation under physiological conditions. These ligands have some promising features in common - they are easily synthesized, contain non-toxic components, have low molecular weights of approximately 300, and are neutral, stable molecules in their biologically active form.

To greatly expand the number of possible hydrogen-bond contacts between ami-nopyrazoles and peptides we created a third generation of hosts relying on oligo-merization of a hitherto unknown unnatural amino acid, 3-aminopyrazole-5-carboxylic acid. This building block can be coupled with itself or other natural and unnatural amino acids by standard peptide synthesis procedures and leads to oligomers with a rigid planar geometry. They have a hydrogen-bond donor and acceptor pattern which can be fine-tuned to be perfectly complementary to that of larger peptides in the ¿-sheet conformation. Because of the five-membered ring,

Fig. 2.4.9. Dimeric 3-aminopyrazole-5-carboxylic acid in its 1:1- and 2:1-complex with a tetrapeptide.

the oligomers have the form of a wide bow, preventing extensive self-association. Modeling experiments suggest that even a tetrapeptide can be bound by two dimeric aminopyrazolecarboxylic acids with a total of 13 hydrogen bonds (Figure 2.4.9). With this increasing number of non-covalent contacts, strong binding should become possible in polar solution, especially with higher aminopyrazole oligomers.

The parent heterocycle can be prepared from the nitro derivative 15 by a variety of reduction techniques. This nitro group, however, can also be conveniently used as a transient protecting group and subjected to standard peptide coupling conditions. It is highly critical to chose the correct protecting group for the pyrazole ring-NH; the p-methoxybenzyl moiety is stable under all the coupling procedures and can be cleaved in the final step with trifluoroacetic acid without causing race-mization of a-amino acids [20].

Hybrid compounds with natural amino acids have a dual advantage over the pure aminopyrazole oligomers. Introduction of amino acids greatly increases the solubility of the whole host compound, and their sequence could serve as affinity tag for recognition of certain characteristic areas on protein surfaces, thereby rendering the 0-sheet ligand specific for a certain protein of interest. A prominent feature for the A0-recognition is, e.g., an internal pentapeptide sequence KLVFF, which is also critical for the aggregation process [21]. For the synthesis of these hybrid systems we developed, in cooperation with the Konig group (Organic Chemistry, Regensburg), a solid phase procedure with Fmoc- and PMB-protected 3-aminopyrazole-5-carboxylic acid as key building block (for more on solid-phase chemistry see Chapters 3.6 and 6.1 and Box 25). Thus, a large variety of aminopyrazole-based oligomers is now accessible by convenient standard proce-

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