Design of Cyclic Peptides - Peptide and Protein Design for Biopharmaceutical Applications - page 155

NMR spectroscopy and the most homogeneous ones were selected. For
example, this technique revealed that the pentapeptide cyclo(- D -Ala-
Ala 4 -) displays a very stable conformation containing a bII 0 -anda
g-turn (see Figure 4.12).
Based on this knowledge, a bioactive sequence, for example ABCDE,
can now be head-to-tail cyclized, where each of the amino acids is used
once in D -configuration. In this way, one obtains five new peptides that
have the same constitution (connectivity) but differ in their stereochem-
istry. Hence, the pharmacophoric side chains are presented in five differ-
ent conformations (see Figure 4.12). The synthesized peptides have
different biological activity, and the bioactive conformation is best repre-
sented in the most active peptide ('spatial screening'). The same proce-
dure can be applied when working with hexapeptides or even larger cyclic
peptides.
4.4.4 General Strategy for Finding Active Hits
As pointed out above, cyclic peptides can exhibit superior properties over
their linear analogues, such as higher activity, better enzymatic stability
and bioavailability, and receptor subtype selectivity. The remaining ques-
tion is how to design cyclic peptides in which these properties are opti-
mized. In this context, the following procedure is suggested:
Step 1: Identification of the target recognition motif. The de novo
approach starts with the identification of molecules that bind to a suitable
target protein. Such molecules can be small peptidic ligands or large
biomacromolecules. To identify the binding sequence, several different
techniques can be used, as mentioned in Section 4.2.1.
Step 2: Elucidation of the pharmacophoric groups. Having found an
attractive binding sequence, an alanine scan [122] is usually performed,
in which each amino acid is systematically substituted by alanine. When
an essential residue is exchanged, a tremendous drop of biological activ-
ity can be observed. For nonessential amino acids, the binding affinity is
affected less drastically, whichmeans that these residues can be optimized
and/or removed.
Step 3: Obtaining initial information of structural arrangements. When the
important amino acids are identified, an initial hint about turn structures is